Characterizations of an ecotype of brake-fern, Pteris vittata, for arsenic tolerance and accumulation in plant biomass
An ecotype of brake fern (Pteris vittata) was assessed for arsenic tolerance and accumulation in its biomass under in vivo and in vitro condition; using soil, and agargelled Murashige and Skoog (MS) medium supplemented with different concentrations of arsenic. The plants were raised in soil amended...
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| Cite this: | Characterizations of an ecotype of brake-fern, Pteris vittata, for arsenic tolerance and accumulation in plant biomass / B.K. Sarangi, T. Chakrabarti // Цитология и генетика. — 2008. — Т. 42, № 5. — С. 16-31. — Бібліогр.: 50 назв. — англ. |
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nasplib_isofts_kiev_ua-123456789-82202025-02-23T18:40:04Z Characterizations of an ecotype of brake-fern, Pteris vittata, for arsenic tolerance and accumulation in plant biomass Вивчення екотипу папороті pteris vittata на стійкість до миш’яку та накопичення рослинної біомаси Изучение экотипа папоротника pteris vittata на устойчивость к мышьяку и накопление растительной биомассы Sarangi, B.K. Chakrabarti, T. Оригинальные работы An ecotype of brake fern (Pteris vittata) was assessed for arsenic tolerance and accumulation in its biomass under in vivo and in vitro condition; using soil, and agargelled Murashige and Skoog (MS) medium supplemented with different concentrations of arsenic. The plants were raised in soil amended with 100–1000 mg arsenic kg^–1 soil, and MS medium was supplemented with 10–300 mg arsenic L–1 medium using Na2HAsO4×7H2O. The spores and haploid gametophytic-prothalli were raised in vitro on MS medium supplemented with arsenic. The field plants showed normal growth and biomass formation in arsenic amended soil, and accumulated 1908– 4700 mg arsenic kg^–1 dry aerial biomass after 10 weeks of growth. Arsenic toxicity was observed above >200 mg arsenic kg^–1 soil. The concentrations of arsenic accumulated in the plant biomass were statistically significant (p < 0.05). Normal plants were developed from spores and gametophyte prothalli on the MS media supplemented with 50–200 mg arsenic L^–1 medium. The in vitro raised plants were tolerant to 300 mg arsenic kg^–1 of soil and accumulated up to 3232 mg arsenic kg^–1 dry aerial biomass that showed better growth performance, biomass generation and arsenic accumulation in comparison to the field plants. Екотип птериса стрічкового (Pteris vittаta) був досліджений на стійкість до миш’яку та його накопичення в біомасі в умовах in vivo і in vitro з використанням грунту та агаризованого середовища МурасігеСкуга (MS), що містять миш’як в різних концентраціях. Рослини вирощували на грунті, що містить 100–1000 мг миш’яку на 1 кг грунту, чи в грунті Мурасіге-Скуга, в котрий додавали 10–300 мг/л Na2HAsO4 ×7H2O. Спори та гаплоїдні гематофітні паростки росли in vitro на середовищі MS з миш’яком. Рослини, які ростуть на грунті, що містить миш’як, характеризувались нормальним ростом і накопиченням біомаси та через 10 тижнів вирощування накопичували 1908–4700 миш’яку на 1 кг сухої надземної біомаси. Токсичність миш’яку проявлялась при його концентрації в грунті більше 200 мг/кг. Концентрації миш’яку, котрі накопичувалися в рослинній біомасі, були статистично значимими (р < 0.5). Зі спор та гаметофітних паростків, котрі вирощували на грунті MS з 50–200 мг/л миш’яку, розвивались нормальні рослини. Отримані in vitro рослини були стійкими до миш’яку в концентрації 300 мг/кг грунту та накопичували миш’як до 3232 мг/кг сухої надземної біомаси, що означає покращання ростових характеристик, формування біомаси та накопичення миш’яку в порівнянні з рослинами, вирощеними на полі. Экотип птериса ленточного (Pteris vittаta) был исследован на устойчивость к мышьяку и его накопление в биомассе в условиях in vivo и in vitro с использованием почвы и агаризованной среды Мурасиге-Скуга (MS), содержащих мышьяк в разных концентрациях. Растения выращивали в почве, содержащей 100–1000 мг мышьяка на 1 кг почвы, или в среде Мурасиге-Скуга, в которую добавляли 10–300 мг/л Na2HAsO4 × 7H2O. Споры и гаплоидные гаметофитные проростки росли in vitro на среде MS с мышьяком. Растения, которые росли в почве, содержащей мышьяк, характеризовались нормальным ростом и накоплением биомассы и через 10 недель выращивания накапливали 1908–4700 мг мышьяка на 1 кг сухой надземной биомассы. Токсичность мышьяка проявлялась при его концентрации в почве свыше 200 мг/кг. Концентрации мышьяка, которые накапливались в растительной биомассе, были статистически значимыми (р < 0,5). Из спор и гаметофитных проростков, которые выращивали на среде MS с 50–200 мг/л мышьяка, развивались нормальные растения. Полученные in vitro растения были устойчивы к мышьяку в концентрации 300 мг/кг почвы и накапливали мышьяк до 3232 мг/кг сухой надземной биомассы, что означает улучшенные ростовые характеристики, формирование биомассы и накопление мышьяка по сравнению с растениями, выращенными в поле. The authors gratefully acknowledge the assistances provided by trainee students for bench works, the instrumentation division of NEERI for arsenic estimation through ICP and Mr. A. Kulkarni, E. B. Division for statistical analysis. The authors are thankful to Director NEERI for providing facility for the work. 2008 Article Characterizations of an ecotype of brake-fern, Pteris vittata, for arsenic tolerance and accumulation in plant biomass / B.K. Sarangi, T. Chakrabarti // Цитология и генетика. — 2008. — Т. 42, № 5. — С. 16-31. — Бібліогр.: 50 назв. — англ. 0564-3783 https://nasplib.isofts.kiev.ua/handle/123456789/8220 en application/pdf Інститут клітинної біології та генетичної інженерії НАН України |
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Оригинальные работы Оригинальные работы |
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Оригинальные работы Оригинальные работы Sarangi, B.K. Chakrabarti, T. Characterizations of an ecotype of brake-fern, Pteris vittata, for arsenic tolerance and accumulation in plant biomass |
| description |
An ecotype of brake fern (Pteris vittata) was assessed for arsenic tolerance and accumulation in its biomass under in vivo and in vitro condition; using soil, and agargelled Murashige and Skoog (MS) medium supplemented with different concentrations of arsenic. The plants were raised in soil amended with 100–1000 mg arsenic kg^–1 soil, and MS medium was supplemented with 10–300 mg arsenic L–1 medium using Na2HAsO4×7H2O. The spores and haploid gametophytic-prothalli were raised in vitro on MS medium supplemented with arsenic. The field plants showed normal growth and biomass formation in arsenic amended soil, and accumulated 1908– 4700 mg arsenic kg^–1 dry aerial biomass after 10 weeks of growth. Arsenic toxicity was observed above >200 mg arsenic kg^–1 soil. The concentrations of arsenic accumulated in the plant biomass were statistically significant (p < 0.05). Normal plants were developed from spores and gametophyte prothalli on the MS media supplemented with 50–200 mg arsenic L^–1 medium. The in vitro raised plants were tolerant to 300 mg arsenic kg^–1 of soil and accumulated up to 3232 mg arsenic kg^–1 dry aerial biomass that showed better growth performance, biomass generation and arsenic accumulation in comparison to the field plants. |
| format |
Article |
| author |
Sarangi, B.K. Chakrabarti, T. |
| author_facet |
Sarangi, B.K. Chakrabarti, T. |
| author_sort |
Sarangi, B.K. |
| title |
Characterizations of an ecotype of brake-fern, Pteris vittata, for arsenic tolerance and accumulation in plant biomass |
| title_short |
Characterizations of an ecotype of brake-fern, Pteris vittata, for arsenic tolerance and accumulation in plant biomass |
| title_full |
Characterizations of an ecotype of brake-fern, Pteris vittata, for arsenic tolerance and accumulation in plant biomass |
| title_fullStr |
Characterizations of an ecotype of brake-fern, Pteris vittata, for arsenic tolerance and accumulation in plant biomass |
| title_full_unstemmed |
Characterizations of an ecotype of brake-fern, Pteris vittata, for arsenic tolerance and accumulation in plant biomass |
| title_sort |
characterizations of an ecotype of brake-fern, pteris vittata, for arsenic tolerance and accumulation in plant biomass |
| publisher |
Інститут клітинної біології та генетичної інженерії НАН України |
| publishDate |
2008 |
| topic_facet |
Оригинальные работы |
| url |
https://nasplib.isofts.kiev.ua/handle/123456789/8220 |
| citation_txt |
Characterizations of an ecotype of brake-fern, Pteris vittata, for arsenic tolerance and accumulation in plant biomass / B.K. Sarangi, T. Chakrabarti // Цитология и генетика. — 2008. — Т. 42, № 5. — С. 16-31. — Бібліогр.: 50 назв. — англ. |
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2025-11-24T11:41:49Z |
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2025-11-24T11:41:49Z |
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1849671820562661376 |
| fulltext |
B.K. SARANGI, T. CHAKRABARTI
National Environmental Engineering Research Institute,
Maharashtra, India,
E.mail: bk_sarangi@neeri.res.in
E.mail: twmneeri@sancharnet.in
CHARACTERIZATION OF AN ECOTYPE
OF BRAKE�FERN, PTERIS VITTATA,
FOR ARSENIC TOLERANCE AND
ACCUMULATION IN PLANT BIOMASS
An ecotype of brake fern (Pteris vittata) was assessed for
arsenic tolerance and accumulation in its biomass under in vivo
and in vitro condition; using soil, and agar�gelled Murashige
and Skoog (MS) medium supplemented with different concen�
trations of arsenic. The plants were raised in soil amended with
100–1000 mg arsenic kg–1 soil, and MS medium was supple�
mented with 10–300 mg arsenic L–1 medium using Na2HAsO4 �
�7H2O. The spores and haploid gametophytic�prothalli were
raised in vitro on MS medium supplemented with arsenic. The
field plants showed normal growth and biomass formation in
arsenic amended soil, and accumulated 1908– 4700 mg arsenic
kg–1 dry aerial biomass after 10 weeks of growth. Arsenic toxici�
ty was observed above >200 mg arsenic kg–1 soil. The concen�
trations of arsenic accumulated in the plant biomass were statis�
tically significant (p < 0.05). Normal plants were developed from
spores and gametophyte prothalli on the MS media supplement�
ed with 50–200 mg arsenic L–1 medium. The in vitro raised
plants were tolerant to 300 mg arsenic kg–1 of soil and accumu�
lated up to 3232 mg arsenic kg–1 dry aerial biomass that showed
better growth performance, biomass generation and arsenic
accumulation in comparison to the field plants.
Introduction. Arsenic is bioactive and potential�
ly toxic. Long�term exposure to low concentrations
of arsenic in drinking water can lead to skin, blad�
der, lung and prostrate cancer, cardiovascular dis�
eases, diabetes, anemia and reproductive, develop�
mental, immunological and neurological effects
[1–4]. Mining and processing of ores of other ele�
ments such as Au, Ag, Cu and Sn in particular had
led to extensive arsenic pollution of mining regions
throughout the world [5–7]. The use of arsenic�
based pesticides as lawn herbicide and insecticides
for rice, orchards and cotton had led to considerable
contamination of domestic and agricultural land [6,
8]. Arsenic contamination in soil is one of the major
sources of arsenic in drinking water [3, 7, 9]. It has
been reported that increased arsenic level in soil
leads to build up of arsenic in plants and crops such
as cereals, vegetables and fruits [10]. Arsenic con�
tamination in soil and water has spread to an alarm�
ing dimension in some eastern states and coastal
parts of India, and other parts in the world [11]. The
remediation of arsenic�contaminated soil and
removal of arsenic from contaminated water is an
important current issue [2]. A number of plants,
known as hyperaccumulators [12], have been iden�
tified accumulating large quantity of contaminants
and a variety of metals in its aboveground biomass
potential for phytoremediation of pollutants [13].
Phytoremediation is an emerging technology that
utilizes the ability of plants to accumulate metals
from soil and groundwater [3, 14–20, 37].
Tolerance to arsenic toxicity is reported in a
large number of plant species, such as Agrostis
tenuis [21], Holcus lanatus [22, 23], Deschampsia
cespitosa and Agrostis capillaris [24], Silene vulgaris
[25, 26], Bidens cynapiifolia [4], Calluna vulgaris
[27], Cytisus striatus [28], Indian mustard [29] and
other plant species [30–32]. Majority of this
arsenic tolerant plants were identified from aban�
doned mine site where the concentration of
arsenic in the soil was extremely high. On the other
hand, plant species like Deschampsia cespitosa [24]
and Silene vulgaris [26] identified from uncontam�
inated soils also exhibited resistance to arsenic.
Under normal conditions, arsenic concentration
in terrestrial plants is less than 10 mg arsenic Kg1
dry biomass [33]. At higher concentrations,
arsenic interferes with plant metabolic processes,
inhibits growth and leads to death. Biomass pro�
duction and crop yield of a variety of species are
significantly reduced at elevated arsenic concen�
trations [34]. The yields of barley (Hordeum vul�
ISSN 0564–3783. Цитология и генетика. 2008. № 516
© B.K. SARANGI, T. CHAKRABARTI, 2008
gare) and rye grass (Lolium perenne) were signifi�
cantly decreased with application of 50 mg arsenic
Kg�1 of soil [10]. The studies on uptake, accumula�
tion and translocation of arsenic in both arsenic�
tolerant and non�tolerant plants have indicated
wide difference in arsenic tolerance among plant
species [21, 23, 36–41].
The brake fern, Pteris vittata (also known as
Chinese brake / ladder brake) is reported as an
arsenic hyperaccumulator plant that could accumu�
late high amount of arsenic in its biomass [36] and
tolerate up to 1500 mg arsenic kg–1 soil in arsenic
amended and contaminated soil. This plant was
identified around California from the South�East of
the USA, and reported to be available in similar
mild climatic areas of the world. Investigations of Tu
and Ma [39] showed that 50 mg arsenic Kg–1 soil
was best for growth and arsenic accumulation in this
fern species, and as high as 2.2 % arsenic was accu�
mulated in the above ground plant biomass and
about 26 % arsenic was removed from the soil. After
8–20 weeks of transplantation, the arsenic biocon�
centration factor (BF) reached 1000 to 1450. The
arsenic translocation factor (TF) in the leaves was
1.2 and 42 at the 2nd week and the 8th week respec�
tively, after of transplantation. Though, Zhang et al.
[40] reported that after 20 weeks of growth in mod�
erately contaminated soil, arsenic translocation and
accumulation in the young parts and old parts of
brake fern was 4893 mg Kg–1 and 7575 mg Kg1
respectively. There are other fern species, which are
also reported to be efficient in accumulating arsenic
in their above ground biomass. Visoottiviseth et al.
[41] assessed the potential of 36 native plant species
of Thailand from mine tailings, where arsenic con�
centration in the soil was up to 16 g Kg–1, and
reported that plant species with the highest leaf
arsenic concentrations did not occur with the high�
est frequency in the contaminated sites. They
reported two species of ferns (Pityrogramma calome�
lanos and Pteris vittata), a herb (Mimosa pudica) and
a shrub (Melastoma malabrathricum) potential for
phytoremediation of arsenic. Francesconi et al. [42]
reported that the silver fern (Pityrogramma calome�
lanos) could accumulate 2760 to 8350 mg arsenic
Kg–1 plant biomass in old fronds, and 5130 to
5610 mg arsenic Kg–1 in young fronds.
The identification of brake fern to be an effi�
cient hyperaccumulator of arsenic has opened
tremendous potentiality for phytoremediation of
arsenic contaminated soil [38, 39]. However,
Meharg [32] and Gumaelius et al. [43] reported that
some members of the genus Pteris; like Pteris
straminea and P. tremula also do not hyperaccumu�
late arsenic, while another species of Pteris cretica
was found to be hyperaccumulator of arsenic.
Gumaelius et al. [43] also investigated arsenic accu�
mulation in the gametophyte and sporophytes of P.
vittata and compared with non�accumulating fern
Ceratopteris richardii. The present work was carried
out to characterize arsenic tolerance and accumula�
tion in an ecotype of Pteris vittata collected from the
Indian subcontinent. Arsenic tolerance of spores,
prothallic�gametophyte and sporophytes were
assessed in arsenic supplemented Murashige and
Skoog [44] growth medium under in vitro condi�
tions, and the sporophytes were assessed in arsenic
supplemented soil under glass house conditions.
Materials and methods. Plant propagation under
glass'house conditions. The ecotype of Pteris vittata
plant was collected from the Kerala state of India,
and used in this study (Fig. 1). The parent plants
were maintained in the glass house in soil pots. The
potting material was a mixture of garden soil, sand
and farmyard manure (FYM) in a ratio of soil : sand :
FYM – 2 : 1 : 1. Young sporophytes were developed
on soil from spores of stock plants, and used in the
experiments. Young plants with 2–3 fronds were
transplanted to single pot with 0.5 kg of soil mix�
ture and allowed to grow for at least one month
prior to experimental use. One to four months old
plants of equal height and with equal number of
fronds were used in the experiments.
ІSSN 0564–3783. Цитология и генетика. 2008. № 5 17
Characterizations of an ecotype of brake'fern, Pteris vittata, for arsenic tolerance and accumulation
Fig. 1. Sporophyte of Pteris vittata ecotype. Bar represents
11.5 cm
Preparation of arsenic amended soil for plant
growth. The soil mixture was amended with differ�
ent concentrations of arsenic for arsenic treatment
experiments by adding Na2HAsO4 · 7H2O to the
soil. The soil mixture was mixed with a basal dose
of fertilizer [N : P : K – 180 : 60 : 120 mg kg–1 soil],
and 500 mL Hoagland nutrient solution [45]
Kg–1 soil mixture to supplement optimum level of
macro and micro nutrients and trace minerals. Two
kg of air�dried soil mixture was amended with ar�
senic to get the desired concentrations of arsenic in
the soil. Calculated amounts of Na2HAsO4 · 7H2O
in aqueous solution were added to the soil mixture
to obtain 50, 100, 200, 500 and 1000 mg total
arsenic Kg–1 soil. The arsenic amended soil was
thoroughly mixed and kept moist for one week
before planting saplings and used for treated
experiments. In a similar manner, soil pots were
prepared without arsenic for raising plants without
arsenic, and used for control experiments.
One to 4 months aged Pteris vittata saplings
were used for the experiments. One healthy fern
plant was transplanted in one packet, amended
with 50/100/200/500/1000 mg arsenic kg–1 soil and
the control was maintained on arsenic free soil.
Five replicates were maintained in each concentra�
tion of arsenic amended soil. The plants were
watered when the topsoil looked dry, and leaching
of water from the soil packets was prevented.
Experimental plants in control and treated soil
were grown inside glass house under similar condi�
tions at an average temperature of 30 °С (day) and
24 °С (night), under a day and night photoperiod
of 14 hours light :10 hours dark period.
The morphological changes of the plants,
appearance of new fronds, coloration of fronds,
and overall growth behavior, were recorded at an
interval of one week, and when noticeable changes
were observed. Live and dead aerial parts of the
plants were collected at different time intervals for
arsenic analysis.
Preparation of arsenic supplemented culture media
for in vitro experimentations under aseptic condition.
Agar gelled Murashige and Skoog (MS) plant tissue
culture medium amended with different concentra�
tions of arsenic was utilized to asses arsenic toler�
ance of the ecotype under in vitro conditions.
Filter�sterilized aqueous Na2HAsO4 · 7H2O solu�
tion was added to molten MS media at <50 °С; to
final concentration of 10, 20, 50, 100 and 200 mg
arsenic L–1 culture medium, inside laminar air�
flow cabinet following standard aseptic procedure.
These media were used in the in vitro experiments
after gelling.
Culture of spore, prothallus, gametophytes and
prothallic�sporophytes. Spores were collected from
the sori of mature fronds from the field grown
sporophytes, decontaminated and cultured on
arsenic supplemented culture media under in vitro
conditions. A piece of leaflet having mature spo�
rangia was wrapped in a packet prepared from fil�
ter paper, and incubated under warm and dry con�
ditions for 3 days. The filter packet with the spores
was dipped in 70 % ethanol for 1 minute followed
by treatment with 0.1 % aqueous mercuric chlo�
ride solution for 3, 5, 7 or 9 minutes and washed
4 times with sterile distilled water. Subsequently,
the filter packet was soaked in sterile water for
7 minutes, and the spores were spread on the sur�
face of the culture medium. In other case, 1cm leaf
cuttings were directly treated with ethanol and
mercuric chloride as described above; the sporan�
gia were ruptured in 0.5ml sterile water and the
entire spore suspension was plated on the agar�
gelled medium. All aseptic works were carried out
under the laminar airflow cabinet. Spores were
plated on agar�gelled MS [44] and Knudson cul�
ture medium [46] without growth hormones. The
in vitro cultures were maintained at 25 ± 2 °С,
1500�lux illumination, 60 % relative humidity and
16 hours light: 8 hours dark photoperiod.
The prothallus, gametophyte and prothallic�
sporophyte that developed from spores, were sub�
cultured on control (without arsenic) and treated MS
medium (supplemented with different concentra�
tions of arsenic). The cultures were incubated under
specified illumination, photoperiod, temperature
and humidity mentioned previously. The spores were
cultured in vitro on full, half and quarter strength
MS, and Knudson hormone free medium supple�
mented with 20–100 mg of arsenic L–1 of culture
medium. The gametophyte prothalli were cultured in
arsenic amended culture medium supplemented
with 100 to 300 mg of arsenic L–1 of medium, and
subcultured to fresh arsenic supplemented medium
at an interval of one month. The gametophytes or
sporophytes were subcultured to fresh medium with
similar or higher concentrations of arsenic, and the
responses of the tissue or organ on the culture medi�
um were monitored from time to time.
ISSN 0564–3783. Цитология и генетика. 2008. № 518
B.K. Sarangi, T. Chakrabarti
Growth of in vitro raised sporophytes in arsenic
amended medium and arsenic amended soil. The
gametophyte prothalli formed 2–4 cm height
sporophyte plants under in vitro, henceforth called
as tissue culture derived plants. These tissue culture
plants were subcultured to fresh arsenic amended
growth medium (100, 200 and 300 mg arsenic L–1)
at an interval of one month and maintained for
6–8 months on the same medium. These sporo�
phytes were subsequently transplanted to soil amend�
ed with the same or higher concentrations of arsenic
and allowed to grow inside glass house conditions.
Arsenic analysis in plant tissue. The live biomass
(green) and dead biomass (dry and brown) were
collected from field plants after 2, 4, 6, and 10
weeks of growth in control and arsenic amended
soil. Likewise live and dead fronds were collected
from the tissue cultured plants, generated in vitro
from spores using arsenic amended culture medi�
um and subsequently transplanted to arsenic
amended soil, when the sporophytes produced suf�
ficient biomass in the arsenic amended soil. These
biomass were thoroughly washed in tap water,
rinsed three times with deionized water, oven dried
at 60 °С for 72 hours and the dry weight was deter�
mined, and used for estimation of arsenic in the
biomass. Known amount of biomass was digested
with analytical grade 6.5ml concentrated HNO3
and 2.5 ml concentrated HCl using the Microwave
Sample Preparation System (Ethos 900, Milestone
Micro�wave Lab. System U.S.) at 300 watt–15
minutes program. The sample solution was filtered
through Whatman filter paper No. 42 and volume
of the filtrate was adjusted to 100 ml with double
distilled water. The concentration of arsenic in the
filtrates was determined by the Inductively Coupled
Plasma spectrometer (ICP�AES, JOBIN YVON,
France) using arsenic standard (MERCK, ICP
standard, product no 1.70303.0100) at concentra�
tions of 0, 1, 5 and 10 mg arsenic L–1 and blank.
Statistical method. The results of effect of soil
arsenic (50 to 1000 mg arsenic kg–1 of soil) on
plant growth at different durations (2–10 weeks),
and arsenic accumulation in the plant biomass
under control and treated conditions were statisti�
cally analyzed. The significance of the differences
in values of arsenic, estimated in plant biomass,
was determined by Pearson Bivariate Correlation
Coefficient and Students t�test, using statistical
software Statistica version 5.1 (Texas, USA).
Results and discussion. Affect of arsenic on
growth and morphology of the sporophytes. Field
grown fern sporophytes (4 months old, with 4–
6 numbers of fronds and about 10 cm height) were
transplanted on control and arsenic amended soil,
with 50, 100, 200, 500 and 1000 mg arsenic kg–1 of
soil. After two weeks, new frond primordia appeared
in control and treated plants (with 50–100 mg
arsenic) whereas, the fronds turned brown in soil
amended with 200 mg arsenic. The fronds of
sporophytes planted in soil >200 mg arsenic kg–1 of
soil dried by two weeks. The symptoms of arsenic
toxicity were first visible in older and larger fronds,
the leaflets in the apical part of fronds started
browning from its tip, and it further extended to
other leaflets and rachis of the affected frond. The
toxic effect began with browning of the leaflets fol�
lowed by drying, and fronds turned black. The
sporophytes transplanted in soil amended with
�500 mg of arsenic kg–1soil dried and turned black
by 5th week. The results of growth performance
indicated a dose dependent cumulative effect of
arsenic toxicity in the plants in comparison to the
control plants (Table 1). Growth and increase in
size of the sporophytes was not affected up to
100 mg arsenic kg–1 soil, and growth response was
similar to the sporophytes raised in control soil
(Fig. 2). New frond primordia appeared after the
second week of transplantation in control as well as
treated soil and the fronds remained partly brown
up to two weeks, but afterwards they enlarged and
turned green like the normal fronds. The height of
the control and treated plants was increased
6–8 cm by the 10th week and the rate of increase in
height was maximum at the 4th week. Whereas, in
case of the plants treated with >200 mg arsenic
kg–1 of soil the increase in plant height was margin�
al, maximum up to 1cm within the same duration
(Fig. 2). It is presumed that the partial browning of
the sporophytes, in control and treated with
100 mg arsenic kg–1 soil, up to the 2nd week could
be due to the transplantation shock to plants in the
soil during acclimatization.
Our findings are comparable with the findings
of Tu and Ma [39], who studied the effect of soil
arsenic on biomass production, arsenic uptake and
accumulation in Pteris vittata identified in
California. In their study, young plants were grown
on arsenic amended soil (950–1500 mg arsenic
Kg–1 soil) in green house, and the highest plant
ІSSN 0564–3783. Цитология и генетика. 2008. № 5 19
Characterizations of an ecotype of brake'fern, Pteris vittata, for arsenic tolerance and accumulation
growth and biomass accumulation (3.9 g plant –1)
were reported with 50 mg arsenic kg–1soil. They
reported arsenic toxicity three days after trans�
planting, the fronds turned dark brown followed by
necrosis in the leaf tips and margins, and plants
died after one week. They reported 64 to 107 %
increase in the fern biomass than control up to
100 mg arsenic kg–1 of soil, and it did not increase
at 200 mg arsenic, whereas at 500 mg arsenic the
above ground biomass was reduced by 64 %. The
reduction in biomass is a common phenomenon of
arsenic phytotoxicity [35]. Tu et al. [38] also
reported slow growth of plants for 6–8 weeks after
transplantation in arsenic soil, though subsequently,
the biomass increased rapidly and nearly quadru�
pled every 4–week.
Arsenic accumulation in aerial biomass of field
plants. The alive and dead fronds of plants, raised
under different concentrations of soil arsenic were
colleted after the 2nd, 4th, 6th and 10th weeks from the
date of plantation from four different sets of plants,
and analyzed to estimate the quantity of arsenic
accumulated in the biomass. The results are pre�
sented in Tables 2 through 5.
The trends of arsenic accumulation in the plant
biomass under different concentrations of soil
ISSN 0564–3783. Цитология и генетика. 2008. № 520
B.K. Sarangi, T. Chakrabarti
Table 1
Morphological changes of Pteris vittata plants grown in control (soil without arsenic) and treated (soil amended
with different concentrations of arsenic) soil
Culture
duration
(in weks)
Control
50 100 200 500 1000
Treated (Arsenic in soil mg kg–1)
No visible change1
2
3
4
5
6
7
8
9
10
Tips and parts of
large, and lower
fronds turned brown
As above
Browning of fronds
increased
One new primordia
initiated
Majority of fronds
(75 %) started brow�
ning
Browning increased
Browning of old
fronds increased
Old fronds dried.
New fronds were
green
Plants were alive as
above
A few leaflets of
larger fronds
turned brown
New frond pri�
mordia appe�
ared
New primordia
developed to
fronds
More new pri�
mordia appe�
ared
Initiation of mo�
re primordia
Primordia deve�
loped to fronds
No visible chan�
ges, plants were
green
More new pri�
mordia develo�
ped
Plant height in�
creased by 6–
8 cm
A few leaflets of
larger fronds tur�
ned brown
New frond pri�
mordia appeared
New primordia
developed to
fronds
More new pri�
mordia appeared
No visible chan�
ges
More primordia
appeared and de�
veloped to fronds
No visible chan
ges, plants were
green
More new pri�
mordia develo�
ped
Plant height in�
creased by 6–
8 cm
A few leaflets of
large fronds
turned brown
New frond pri�
mordia appe�
ared
New primordia
developed to
fronds
As above, brown
fronds dried
More new frond
primordia initi�
ated and devel�
oped
No visible chan�
ges
Upper part of
fronds (10 %)
browned
No change.
Plants were gre�
en
Plant height in�
creased by 6–
8 cm
Peripheral parts of
large fronds turned
brown
Venation of the
fronds were visibly
brown
As above, new
frond primordia
initiated
Brown fronds died
More fronds tur�
ned brown
All fronds turned
brown
Plant turned brown
and dried
–
–
Many fronds
turned brown
Brown fronds
dried
More fronds
turned brown
Browning of
fronds increa�
sed
More fronds
turned com�
pletely brown
Brown fronds
died. All fronds
started brow�
ning
Plant turned
brown and dri�
ed
–
–
arsenic after 2,4,6 and 10 weeks of growth were
analyzed (Fig. 3). The arsenic accumulation in the
plant biomass indicates that accumulation was more
at higher levels of soil arsenic, and when duration of
plant growth was more. Thus, arsenic accumulation
in this ecotype of brake fern varied in accordance to
the level of arsenic in the soil and duration of plant
growth. The amount of arsenic accumulated in
plant biomass (mg of arsenic kg–1 dry weight) grown
in the soil amended with 100 and 200 mg arsenic
kg–1 soil was comparable to that with 500 and
1000 mg arsenic kg–1 soil (Fig. 3). Whereas, in case
of the plants grown with similar concentration of
soil arsenic, arsenic accumulation in plant biomass
was quantitatively more after ten weeks than the
plants grown for less duration (Fig. 4).
The analyses indicate that, the rate of arsenic
transportation to the shoot in arsenic amended soil
was less in all concentrations of arsenic up to 4 weeks
of plant growth, which, however, increased sharply
between 6 and 10 weeks (Fig. 5). This ecotype of
ІSSN 0564–3783. Цитология и генетика. 2008. № 5 21
Characterizations of an ecotype of brake'fern, Pteris vittata, for arsenic tolerance and accumulation
Fig. 2. Growth performance of field grown sporophytes of P. vittata in control and As spiked soil after 8 weeks (A – control,
B – 50 mg As, C – 100 mg As, D – 200 mg As Kg–1 soil) and 10 weeks (E – control, F – 50 mg As, G – 100 mg As, H – 200 mg
As Kg–1soil). Bar represents 10 cm
Fig. 3. Correlation of quantity of arsenic accumulated in bio�
mass (dry weight) of the Pteris vittata ecotype with respect to
different levels of soil arsenic; 50, 100, 200, 500 and 1000 mg
arsenic kg–1 of soil after 2, 4, 6 and 10 weeks of growth
brake fern accumulated 1908 mg arsenic kg–1 of dry
biomass when grown in arsenic amended soil for a
period of 10 weeks with lower concentrations of
arsenic (50–100 mg arsenic kg–1 soil). Whereas, at
higher concentrations of arsenic (500–1000 mg
arsenic kg–1 soil), in the same growth period, more
quantity of arsenic (4700 mg arsenic kg–1 of dry plant
biomass) was accumulated in the plant biomass.
Moreover, under high concentrations of soil arsenic
(1000 mg arsenic kg–1 soil), very high quantity of
arsenic was accumulated in the plant biomass as
early as after 2 weeks (Fig. 4). However, under 1000
mg As the increase in plant biomass was negligible
and aerial parts of the plants gradually turned black
and subsequently the plants died. The variances in
arsenic concentrations in the plant biomass at differ�
ent soil arsenic concentrations (control vs1000 mg,
50 vs1000 mg, 100 vs1000 mg, 200 vs100 mg,
200 vs500 mg and 500 vs1000 mg arsenic kg–1 soil),
and at different duration of growth (2nd vs10th, 4th vs
10th and 6th vs10th week) were statistically analyzed.
The differences were significant at 95 % confidence
level (*p < 0.05).
Arsenic accumulated in dead and live fronds of
this fern ecotype was estimated to determine the
rate of arsenic accumulation in old and young aer�
ial parts. The concentrations of arsenic in the aer�
ial biomass after 2 weeks and 4 weeks of growth in
arsenic amended soil are presented in Table 6. The
results show that the rate of arsenic accumulation
was almost similar between live and dead fronds
after 2 and 4 weeks of growth at all concentrations
of soil arsenic, indicating that the rate of arsenic
translocation to the aerial parts was almost similar
in all fronds. The comparative rate of arsenic accu�
mulation after the 2nd and the 4th weeks of growth
with relation to all concentrations of soil arsenic
was analyzed (Fig. 6). It was found that larger
amount of arsenic was deposited in dead biomass
of plants than that of live biomass in case of plants
grown up to 4 weeks. This could be because the
plants grown for 4 weeks in arsenic amended soil
were exposed to arsenic for more duration, which
led to accumulation of more amount of arsenic in
plant biomass in comparison to the plants grown in
the arsenic amended soil for 2 weeks.
The study indicates that this ecotype of brake
fern is efficient in extracting arsenic form the soil
up to 100 mg arsenic kg–1 soil, and can tolerate up
to 200 mg arsenic kg–1 soil. This genotype of the
brake fern accumulated 1908 mg arsenic kg–1 dry
biomass in lower concentrations of soil arsenic
(100 mg arsenic kg–1 soil) after 10 weeks of growth.
ISSN 0564–3783. Цитология и генетика. 2008. № 522
B.K. Sarangi, T. Chakrabarti
Table 2
Plant height, biomass and quantity of arsenic accumulated
in plant biomass of the Pteris vittata ecotype after 2 weeks
of arsenic treatment
Concentration
of arsenic in
soil (mg kg–1)
Height of plant
(cm, ±SD)
Fresh weight
of plant
(g, ±SD)
Quantity of
arsenic in plant
biomass after 2
weeks (mg kg–1)
Control
50
100
200
500
1000
19.5 ± 2.5
14.5 ± 2.0
22 ± 2.1
15 ± 1.5
26.5 ± 3.5
17.5 ± 1.1
4.47 ± 1.2
4.86 ± 1.0
3.64 ± 0.5
4.54 ± 0.8
2.25 ± 0.9
4.46 ± 1.5
14
54
164
216.46
874
2274
Concentration
of arsenic in
soil (mg kg–1)
Height of plant
(cm, ±SD)
Fresh weight
of plant
(g, ±SD)
Quantity of
arsenic in plant
biomass after 4
weeks (mg kg–1)
Control
50
100
200
500
1000
17 ± 0.5
21 ± 1.5
19,5 ± 2.5
15 ± 1.9
12 ± 0.5
14.5 ± 1.3
5.728 ± 0.5
14.94 ± 4.5
8.94 ± 2.1
6.26 ± 0.5
7.84 ± 2.5
3.9 ± 0.5
27
46.54
165.08
206.25
720
3308
Concentration
of arsenic in
soil (mg kg–1)
Height of plant
(cm, ±SD)
Fresh weight
of plant
(g, ±SD)
Quantity of
arsenic in plant
biomass after 6
weeks (mg kg–1)
Control
50
100
200
500
1000
21 ± 1.5
9 ± 0.5
24 ± 1.5
12.5 ± 0.5
11.5 ± 2.5
12 ± 3.5
12.05 ± 0.5
1.09 ± 0.5
15.79 ± 3.5
2.39 ± 0.9
1.46 ± 0.7
1.23 ± 0.9
30.00
198.29
191.35
637.89
2573.67
4106.41
Table 3
Plant height, biomass and quantity of arsenic accumulated
in plant biomass of the Pteris vittata ecotype after 4 weeks
of arsenic treatment
Table 4
Plant height, biomass and quantity of arsenic accumulated
in plant biomass of the Pteris vittata ecotype after 6 weeks
of arsenic treatment
Under high soil arsenic (1000 mg kg–1 soil), the
plant biomass accumulated as high as 4700 mg
arsenic kg–1 dry weight, in the same 10 weeks growth
period. Further, under high concentration of soil
arsenic (1000 mg kg–1 soil) after 2 weeks of growth,
this genotype also accumulated high amount of
arsenic (2300 to 2428 mg kg–1 dry weight) in the
plant biomass.
Our findings on arsenic accumulation by this
ecotype of Pteris vittata are comparable to the Pteris
vittata ecotype reported by Tu and Ma [39]. The
brake fern was reported to be a hyperaccumulator of
arsenic by Ma et al. [36]. This species was identified
at California, which accumulated up to 15,861 mg
arsenic kg–1 plant biomass in the aerial shoots after
2 to 6 weeks and arsenic toxicity was severe
�500 mg arsenic Kg–1 in soil. However, in the same
species of brake fern (ladder brake) Tu and Ma [39]
reported that arsenic accumulation was maximum
in the young plants under 50 mg arsenic Kg–1 soil,
based on the results after 12 to 18 weeks of growth
of the plants with 50–1500 mg arsenic Kg–1 soil in
green house conditions. Under low concentrations
of soil arsenic, the younger fronds accumulated high
concentration of arsenic, whereas, at high arsenic
concentration in soil accumulation was more in the
older fronds, presumably, due to older fronds
receiving arsenic for a longer time in comparison to
the younger fronds. Further study on Pteris vittata
indicated that arsenic accumulation in the fronds
increased with duration of growth [38]. The mature
and young fronds had 6610 and 5570 mg arsenic
Kg–1 biomass respectively and arsenic concentra�
tions were highest in old fronds (13 800 mg Kg–1 in
dry biomass). In the same ecotype of Pteris vittata
Zhang et al. [40] reported that arsenic accumula�
tion was 4893 mg Kg–1 and 7575 mg Kg–1 plant bio�
mass in young and old parts of the plants respective�
ly, after 20 weeks of growth. Concentration of
arsenic was lowest in the roots of plants, substantial�
ly high in fronds, and old fronds had highest con�
centrations of arsenic. Arsenic accumulation by this
ecotype of Pteris vittata from the Indian subconti�
nent is less in comparison to the Pteris vittata eco�
type identified at California [36], but our results
with this ecotype are similar to the findings report�
ed for Pteris vittata by other workers [38–40].
Ma et al. [36] estimated that the bioaccumula�
tion factor (BF) for arsenic by brake fern was as
high as 193 as an indicator of efficient arsenic accu�
ІSSN 0564–3783. Цитология и генетика. 2008. № 5 23
Characterizations of an ecotype of brake'fern, Pteris vittata, for arsenic tolerance and accumulation
Fig. 4. Comparison of quantity of arsenic accumulated in
plant biomass (dry weight) of the Pteris vittata ecotype after
2 weeks, 4 weeks, 6 weeks and 10 weeks of growth at different
levels of soil arsenic (50, 100, 200, 500 and 1000 mg arsenic
kg–1 soil)
Fig. 5. Rate of arsenic accumulation in the ecotype of Pteris
vittata after 2, 4, 6 and 10 weeks at different concentrations
of soil arsenic (50–1000 mg arsenic kg–1 soil)
Fig. 6. Quantity of arsenic accumulated in live and dead bio�
mass of Pteris vittata ecotype treated with different concen�
trations of arsenic amended in soil, after two and four weeks
mulation in plant biomass from soil. While, Tu et al.
[38] reported that the BF for arsenic accumulation
in the fronds of Pteris vittata based on water�solu�
ble arsenic was increased to >1000 in the fronds
after 8 week of transplanting. In this ecotype of
brake fern studied by us, the difference in arsenic
accumulation between live and dead fronds was
marginal after two weeks of growth in arsenic
amended soil (100 and 200 mg arsenic kg–1 soil).
However, after 4 weeks of growth in arsenic
amended soil, the difference in arsenic accumula�
tion was significant between dead and live fronds
(Table 6). Tu and Ma [39] have also reported that
Pteris vittata had highest BF value for arsenic with
50 mg arsenic Kg–1 soil. In this ecotype of brake
fern, the bio�concentration factor of arsenic in the
plant biomass was up to 19 at 100 mg arsenic Kg–1
soil after 10th week (Table 5). The BF for arsenic
varied with respect to different concentrations of
soil arsenic and duration of growth.
Growth performance of the sporophytes developed
from spores on arsenic supplemented MS medium in
vitro and arsenic amended soil. Spores of the Pteris
vittata plant were aseptically cultured on full, half
and quarter strength Murashige and Skoog and
Knudson hormone free medium supplemented
with various concentrations of arsenic to assess tol�
erance to arsenic toxicity. The spores germinated
gametophyte prothalli in all media, but development
of sporophytes from the gametophytes was better
supported in full strength MS medium (Fig. 7,
B–D). The gametophyte prothalli were cultured in
arsenic amended MS medium with different con�
centrations of arsenic to assess arsenic tolerance at
organ level. After 2–3 months, the gametophytic�
prothalli generated fern sporophytes on control MS
medium, as well as arsenic treated medium supple�
mented with 20–100 mg arsenic L–1 of medium
(Fig. 8, C–F). The tender sporophytes were mor�
phologically normal, and did not show visible
symptoms of arsenic toxicity. These prothallic
gametophytes and sporophytes were further cul�
tured on higher concentrations of arsenic (Fig. 9
and 10), up to 300 mg arsenic L–1 of medium.
These tissue culture derived sporophytes were
separated from the gametophytic�prothalli after 2–
4 cm height and transplanted on fresh growth medi�
um amended with 100, 200 and 300 mg arsenic L–1
of medium (Fig. 10). They were subcultured to the
same medium at an interval of one month and
maintained under in vitro conditions for 6–8
months. These plants developed intricate root sys�
tems, height and the above ground biomass of the
plants increased to more than double after ten we�
eks. Under in vitro conditions with 200 mg arsenic
L–1 of growth medium, sporophytes were similar to
control plants. The sporophytes treated with
300 mg arsenic L–1 MS medium had less biomass in
comparison to the plants treated with 200 mg ar�
senic L–1 of medium (Fig. 10); however, the plants
were green and alive after repeated subculture to ar
senic supplemented medium with 300 mg arsenic
L–1 of medium. Observations of growth response of
these sporophytes in arsenic amended growth
medium up to 8 months are presented in Table 7.
Subsequently, these in vitro raised sporophytes
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B.K. Sarangi, T. Chakrabarti
Concentration
of arsenic in
soil (mg kg–1)
Height of plant
(cm)
Fresh weight
of plant (g)
Quantity of
arsenic in plant
biomass after 6
weeks (mg kg–1)
Control
50
100
200
500
1000
17 ± 3.5
21 ± 1.5
19,5 ± 0.5
15 ± 1.5
12 ± 3.5
14,5 ± 3.0
5.72 ± 0.5
14.94 ± 1
8.945 ± 1.5
6.26 ± 0.3
7.84 ± 2.5
3.9 ± 0.7
48.89
347.54
1908.53
1827
3593
4700
Table 5
Plant height, biomass and quantity of arsenic accumulated
in plant biomass of Pteris vittata ecotype after 10 weeks
of arsenic treatment
Table 6
Quantity of arsenic accumulated in the live and dead
biomass of control and arsenic treated Pteris vittata
ecotype after 2 weeks and 4 weeks, mg kg–1
Concentration
of arsenic
in soil
Arsenic
in live
biomass
Arsenic
in dead
biomass
Arsenic
in live
biomass
Arsenic
in dead
biomass
After the 2nd week After the 4th week
Control
50
100
200
500
1000
14
54
190
242
900
2300
Dead
fronds
absent
620
122
309
516
2428
27
39
158
199
713
3301
Dead
fronds
absent
386
296
404
477
4736
were transplanted on arsenic amended soil and
raised under glass house.
Performances of in vitro raised Pteris vittata
sporophytes after transplantation to arsenic amended
soil. The Pteris vittata sporophytes; 8–10 cm in
height, with 10–20 fronds and six to eight months
old, which were developed in vitro on arsenic (150,
200 and 300 mg L–1) amended MS media, were
transplanted on arsenic amended soil (150, 200,
300 and 500 mg kg–1 soil) and grown in glass
house. The sporophytes selected with150 and 200
mg arsenic L–1 MS media were transplanted to soil
amended with the same and higher (300 and 500
mg arsenic kg–1 soil) concentrations of arsenic. It
was observed that after transplantation to arsenic
amended soil these sporophytes established in less
time, in comparison to the sporophytes developed
ex vitro in soil. A few fronds started browning in
control (soil not amended with arsenic) as well as
in treated plants (soil amended with different con�
centrations of arsenic), probably due to the trans�
plantation shock. However, both in control and
treated plants new frond primordia emerged
quickly by the 4th week, and enlarged to normal
sized fronds. The length of new fronds increased to
�10 cm after 6 weeks, the leaflets were long, and
all plants were green and healthy. In case of the
plants treated with 300 mg arsenic kg–1 of soil, the
new fronds looked brown up to the 4th week, but,
subsequently the fronds in control and treated
plants looked alike. The growth rate of sporophyte
plants in 100 and 200 mg arsenic kg–1 soil was nor�
mal (Fig. 11), whereas the growth rate was slow in
300 mg arsenic kg–1 soil. All fronds of control
plants and treated with 100, 200 and 300 mg
arsenic kg–1 of soil were enlarged in length to
�20 cm by the 9th week and the numbers of fronds
were �20. In case of the plants treated with 300 mg
arsenic kg–1 soil, the lower 2–3 fronds of the plant
developed black patches on the peripheral parts of
leaflets and adjoining the venation of the frond,
but growth performances of the treated plants were
similar to control plants. Whereas, the height of
the sporophytes and size of the fronds did not
increase in soil treated with 500 mg arsenic kg–1 of
soil, the fronds turned black and the sporophytes
degenerated.
The in vitro generated Pteris vittata sporophytes
grew better with 200 and 300 mg arsenic kg–1 soil
after transplantation to arsenic amended soil
ІSSN 0564–3783. Цитология и генетика. 2008. № 5 25
Characterizations of an ecotype of brake'fern, Pteris vittata, for arsenic tolerance and accumulation
Fig. 7. Spore culture: development of gametophytes (A –
Knudson medium, B – MS medium), and sporophytes (C,
D – microscopic view of nascent sporophytes) on arsenic fee
agar�gelled media after 4–6 weeks. Bar represents 13 mm
(A, B, C),1.0 mm (D)
Fig. 8. Growth responses of gametophyte prothalli of Pteris
vittata in MS agar�gelled media: without arsenic (A – single
gametophyte prothalli microscopic view; B – clump of
gametophyte), and supplemented with different concentra�
tions of arsenic (C – 20 mg, D – 60 mg, E – 100 mg and F –
200 mg As L–1) after 6–9 weeks. Shows development of spo�
rophytes. Bar represents 0.25 mm (A), 13 mm (B–F)
(Table 8), as compared to the sporophytes raised in
soil and treated with similar concentrations of
arsenic amended soil (Table 1). It was also observed
that the size of plants, number of fronds and gener�
ation of biomass were more in case of the in vitro
raised plants (Fig. 11) in comparison to plants gen�
erated in ex vitro condition and transplanted in
arsenic amended soil (Fig. 2). This could be
because the in vitro raised plants were treated with
arsenic from the very early stage of growth, and the
plants were adapted to tolerate higher concentra�
tions of arsenic when transplanted to arsenic
amended soil under field conditions. Presumably,
as a result of arsenic induced enhanced expression
of genetic traits and physiological adaptations of
the in vitro raised plants due to prolonged arsenic
treatment from early stage of growth and develop�
ment in vitro. Further biochemical characteriza�
tion of in vitro raised and field raised arsenic treat�
ed plants and investigations with genetic markers
specific for arsenic tolerance would justify for this
difference in their nature.
Arsenic accumulation in the biomass of in vitro
raised sporophytes after 12 months growth on arsenic
amended soil. The in vitro developed plants grew
luxuriantly in 100, 200 and 300 mg arsenic kg–1 of
soil, and generated normal biomass by twelve
months (Table 8 and Fig. 11). The average num�
bers of live and dead fronds in twelve months old
plants didnґt show significant difference between
control and arsenic treated plants. The concentra�
tions of arsenic accumulated in the live and dead
fronds of these plants were estimated using
Inductively Coupled Plasma (ICP) Spectrophoto�
meter as described earlier under the Materials and
Methods. The amounts of arsenic, in live and dead
fronds of plants (mg of arsenic kg–1 of dry weight,
Table 8) indicated greater accumulation of arsenic
in the live fronds in comparison to the dead fronds
in all concentrations of arsenic (100, 200, 300 and
500 mg arsenic kg–1 of soil). It would seem that
arsenic was actively transported to the live fronds
that were in a physiologically active state of growth
in comparison to the dead fronds. The maximum
concentration of arsenic in live frond of plants was
3232 mg kg–1 dry weight, which were grown under
300 mg arsenic kg–1 of soil for duration of
12 months. The plants grown under 500 mg
arsenic kg–1 of soil had a very few fronds, but some
of them were still green. The live and dead fronds
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B.K. Sarangi, T. Chakrabarti
Fig. 9. Profuse growth of
gametophytes (A) and
emergence of sporophytes
from the gametophytes (B,
C) in MS medium supple�
mented with 50 mg (A),
100 mg (B) and 150 mg
(C) As L–1 medium. Bar
represents 11.6 mm
Fig. 10. Growth perform�
ances of in vitro developed
sporophytes in MS medium
supplemented with 100 mg
(A), 200 mg (B) and 300 mg
(C) arsenic L–1 of medium.
Bar represents 11.6 mm
of these plants treated with 500 mg arsenic had
3732 mg and 3237 mg arsenic kg–1 dry biomass
respectively. The observations indicated that the
estimated concentration of arsenic in the live
fronds of this ecotype of Pteris vittata induced toxic
effect and could be the threshold concentrations
within the accumulated plant biomass.
The Pteris vittata plants collected by Ma et al.
[36] accumulated 11.8–64 mg arsenic kg–1 above
ground biomass in uncontaminated soil (arsenic le�
vel: 0.47–7.56 mg arsenic kg–1 soil), and up to 1400–
7500 mg arsenic kg–1 biomass in arsenic contaminat�
ed soil (arsenic level: 18.8–1603 mg arsenic kg–1
soil). Our investigations with this Indian ecotype of
brake�fern also indicate similar observations within
the study period of 10 weeks. The growth character�
istics and biomass accretion of that plant in arsenic
contaminated soil indicated that biomass increase
took place after a slow growth for 6–8 weeks [38],
arsenic accumulation in the fronds increased with
growth of the plant that was a function of time, and
highest concentrations of arsenic was in old fronds.
Gumaelius et al. [43] reported that gametophytes of
P. vittata grow normally in the medium containing
20 mM arsenate, and accumulate >2.5 % of their
dry weight as arsenic in a manner similar to that the
ІSSN 0564–3783. Цитология и генетика. 2008. № 5 27
Characterizations of an ecotype of brake'fern, Pteris vittata, for arsenic tolerance and accumulation
Table 7
In vitro growth response of the Pteris vittata sporophytes in arsenic amended MS media supplemented
with 50, 100 and 200 mg arsenic L–1 medium
Type of Culture
2 5
Ht – 4, Fnd – 10,
4–5, Lft – 1–2,
green, Rt – 2–3
Ht – 1.5, Fnd – 5,
2, Lft – Small,
brownish, Rt –
Slender
Ht – 4.5, Fnd –
7–8, 4, Lft –
Enlarged, green,
Rt – Several, elon�
gated, brown
Ht – 3 cm, Fnd –
4, 3 cm, Lft –
Small, green, Rt –
Diminutive
Ht – 5, Fnd – 10,
4–5, Lft – 2, green,
Rt – Several,
brownish
Ht – 2, Fnd – 6, 2,
Lft – Small, Rt –
Few
Ht – 4.5, Fnd – 7–
8, 6.5, Lft – Large
and green, Rt –
Numerous, intricate
Ht – 3 cm, Fnd –
4, 3 cm, Lft –
Small, green, Rt –
Meager
Ht – 8, Fnd –
12–14, 6–7, Lft –
3–5, green, Rt –
Numerous
Ht – 2, Fnd – 8, 3,
Lft – Small, Rt –
Scarce
Ht – 7.3, Fnd – 10,
7, Lft – Large and
green, Rt –
Elongated, blackish
Ht – 3 cm, Fnd –
5, 4 cm, Lft –
Small, green, Rt –
Scarce
Ht – �15, Fnd –
30, 12, Lft – 2–4,
green, Rt – Profuse,
intricate, brown to
black, + –50 times
Ht – �15, Fnd –
28, 10, Lft – 2–
4 cm, green, Rt –
Profuse, intricate,
brown to black,
+ –50 times
Ht – �15, Fnd –
30, 10, Lft – 2–4
green, Rt – Profuse,
intricate, brown to
black, + –50 times
Ht – 8 cm, Fnd – 8,
5 cm, Lft – 1–2,
green, Rt – Scarce,
+ –15 times
7 10 32
Growth response, weeks
MS medium
Control
MSAs150
MSAs200
MSAs300
Ht – 2, Fnd – 7,
2–3, Lft – 1, green�
ish brown, Rt –
Short
Ht – 1.5, Fnd – 4,
1–2, Lft – Small,
brownish, Rt – Not
seen
Ht – 3, Fnd – 5,
2–3, Lft – Small,
green, Rt – Short
Ht – 3 cm, Fnd –
4, 3 cm, Lft –
Small, green, Rt –
Not seen
Note. As – mg arsenic L–1 MS medium; Ht – height of plants in cm; � – to the size of culture container; Fnd – no of
fronds and length in cm; Lft – size of leaflets in cm, and color; Rt – number of roots, size and color; + – increase in plant
size from initial size.
Fig. 11. Growth performance of in vitro developed sporo�
phytes of P. vittata in As spiked treated soil (A–100 mg K–1,
B–200 mg Kg–1) after 12 months. Bar represents 10.2 cm
sporophytes. Whereas, the gametophytes of the
related non�accumulating fern Ceratopteris richardii
die at even low (0.1 mM) arsenic concentrations,
and the gametophytes of the relatedarsenic accumu�
lator Pityrogramma calomelanos tolerate and accu�
mulate arsenic to intermediate levels compared to P.
vittataand C. richardii. Their study also revealed nat�
ural variability in arsenic tolerance in the gameto�
phyte populations from 40 different P. vittata sporo�
phyte plants collected at different sites in Florida.
The spores and prothallic�gametophyte of this eco�
type of Pteris vittata grow in MS culture medium in
vitro with 20–100 mg/L and up to 300 mg/L arsenic
respectively. Germination of spores, growth per�
formance of prothallic�gametophytes as well as
development of saprophytes from the gametophytes
was normal under arsenic supplemented growth
medium in vitro.
Conclusion. Arsenic tolerance in plants may
result from arsenic exclusion through avoidance or
restriction of arsenic uptake and transport to the
shoots [31, 47] or accumulation of higher concen�
tration of arsenic within plant tissue in comparison
to the surroundings [15,18, 31]. The molecular
analysis of arsenic metabolism in terrestrial plant
species have shown that arsenate�induced
hytochelatins (PC) accumulation and PC�based
arsenic sequestration are responsible for both nor�
mal and enhanced arsenate tolerance [30, 48–50].
The results of arsenic tolerance and accumula�
tion by this ecotype of Pteris vittata used in this
study show that arsenic accumulation in the plant
biomass was statistically significant (*p < 0.05) at
all concentrations of soil arsenic (50, 100, 200,
500 and 1000 mg kg–1 soil), and at different dura�
tions (2nd –10th week, 4th–10th week and 6th–10th
week). The in vitro raised Pteris vittata plants were
tolerant to higher concentrations of arsenic
(300 mg arsenic kg–1 of soil) in comparison to the
plants raised in ex vitro conditions (field grown
plants) using NaH2AsO4 · 7H2O as the source of
arsenic, which is more toxic to plants in compari�
son to its potassium salt. The growth performanc�
es of field grown plants were similar to control
plants at 100 mg arsenic Kg–1 soil. But, the field
grown plants generated poor biomass under
�200 mg arsenic kg–1 soil and the fronds turned
brown between 3–6 weeks (Table 1). However, at
this concentration of soil arsenic, the field plants
accumulated 216 and 1827 mg arsenic kg–1 dry
biomass of fronds after 2 and 10 weeks of treat�
ment respectively (Table 2 and 5). On the contrary,
the in vitro raised plants showed normal growth
performance and biomass accumulation in 100,
200 and 300 mg arsenic kg–1 soil, which was better
than the field plants (Table 7 and 8). Under high
concentrations of soil arsenic (300 mg arsenic kg–1
soil), the in vitro selected plants accumulated
3232 mg arsenic and 2132 mg arsenic kg–1 live and
dead frond biomass respectively (Table 8). Arsenic
accumulation was more in live as well as dead
fronds in case of in vitro raised plants (Table 8) in
ISSN 0564–3783. Цитология и генетика. 2008. № 528
B.K. Sarangi, T. Chakrabarti
Table 8
Morphological features and quantity of arsenic accumulated in the aerial biomass of in vitro developed Pteris vittata
sporophytes after growth in arsenic amended soil for 12 months
Concentration of arsenic
mg kg–1 soil
Height
of plant (cm)
Average length
of fronds (cm)
Live Dead Live Dead
Concentration of arsenic in plant
biomass (mg kg–1 of dry weight)
Average number of fronds
Control (without arsenic)
As150
As200
As300
As500
34 ± 5
34 ± 5
32 ± 3
32 ± 5
8 ± 5
32 ± 7
30 ± 2
32 ± 5
30 ± 5
9 ± 5
26 ± 3
25 ± 2
28 ± 5
28 ± 5
2
7�dry
3�partly dry
7�dry
3�partly dry
8�dry
4�partly dry
8�dry
4�partly dry
8�dry
24.5
1751
2046.5
3232
3731.5
17
951.5
1326.5
2132
3236.5
Note. As – mg arsenic kg–1 of soil.
comparison to the field raised plants (Table 6). The
in vitro raised Pteris vittata plants were more toler�
ant to arsenic stress and accumulated more arsenic
in the biomass. The mechanism of arsenic hyper
accumulation in the plant biomass of Pteris vittata
could be through compartmentalization or by
molecular chaperons such as phytochelatins,
which is being investigated.
This ecotype of brake�fern identified is hardy,
perennial and survives under high temperature at
40–45 °С (data not presented), and adapt to new
environment easily. It could be useful for phytoex�
traction of arsenic from contaminated soil. Further
studies are carried out to assess genetic variations
between the ecotypes of Pteris vittata plants for
arsenic uptake and metabolism, and find out the
reason for better survival and tolerance of in vitro
plants in comparison to the field plants under high�
er concentrations of arsenic. This could help in
selection of brake fern species and clones showing
better arsenic hyper accumulation and suited for
remediation of arsenic contaminated soils.
The authors gratefully acknowledge the assis�
tances provided by trainee students for bench works,
the instrumentation division of NEERI for arsenic
estimation through ICP and Mr. A. Kulkarni, E. B.
Division for statistical analysis. The authors are
thankful to Director NEERI for providing facility for
the work.
Б.К. Саранги, Т. Чакрабарти
ИЗУЧЕНИЕ ЭКОТИПА ПАПОРОТНИКА
PTERIS VITTATA НА УСТОЙЧИВОСТЬ К
МЫШЬЯКУ И НАКОПЛЕНИЕ
РАСТИТЕЛЬНОЙ БИОМАССЫ
Экотип птериса ленточного (Pteris vittаta) был ис�
следован на устойчивость к мышьяку и его накопление
в биомассе в условиях in vivo и in vitro с использова�
нием почвы и агаризованной среды Мурасиге�Скуга
(MS), содержащих мышьяк в разных концентрациях.
Растения выращивали в почве, содержащей 100–
1000 мг мышьяка на 1 кг почвы, или в среде Мурасиге�
Скуга, в которую добавляли 10–300 мг/л Na2HAsO4 �
� 7H2O. Споры и гаплоидные гаметофитные про�
ростки росли in vitro на среде MS с мышьяком. Рас�
тения, которые росли в почве, содержащей мышьяк,
характеризовались нормальным ростом и накопле�
нием биомассы и через 10 недель выращивания на�
капливали 1908–4700 мг мышьяка на 1 кг сухой
надземной биомассы. Токсичность мышьяка прояв�
лялась при его концентрации в почве свыше 200 мг/кг.
Концентрации мышьяка, которые накапливались в
растительной биомассе, были статистически значи�
мыми (р � 0.5). Из спор и гаметофитных проростков,
которые выращивали на среде MS с 50–200 мг/л мы�
шьяка, развивались нормальные растения. Полученные
in vitro растения были устойчивы к мышьяку в кон�
центрации 300 мг/кг почвы и накапливали мышьяк до
3232 мг/кг сухой надземной биомассы, что означает
улучшенные ростовые характеристики, формирова�
ние биомассы и накопление мышьяка по сравнению с
растениями, выращенными в поле.
Б.К. Сарангі, Т. Чакрабарті
ВИВЧЕННЯ ЕКОТИПУ ПАПОРОТІ
PTERIS VITTATA НА СТІЙКІСТЬ
ДО МИШ’ЯКУ ТА НАКОПИЧЕННЯ
РОСЛИННОЇ БІОМАСИ
Екотип птериса стрічкового (Pteris vittаta) був до�
сліджений на стійкість до миш’яку та його накопи�
чення в біомасі в умовах in vivo і in vitro з викорис�
танням грунту та агаризованого середовища Мурасіге�
Скуга (MS), що містять миш’як в різних концентраціях.
Рослини вирощували на грунті, що містить 100–1000 мг
миш’яку на 1 кг грунту, чи в грунті Мурасіге�Скуга, в
котрий додавали 10–300 мг/л Na2HAsO4 · 7H2O. Спори
та гаплоїдні гематофітні паростки росли in vitro на се�
редовищі MS з миш’яком. Рослини, які ростуть на
грунті, що містить миш’як, характеризувались нор�
мальним ростом і накопиченням біомаси та через 10
тижнів вирощування накопичували 1908–4700 миш’я�
ку на 1 кг сухої надземної біомаси. Токсичність ми�
ш’яку проявлялась при його концентрації в грунті
більше 200 мг/кг. Концентрації миш’яку, котрі на�
копичувалися в рослинній біомасі, були статистично
значимими (р � 0.5). Зі спор та гаметофітних парост�
ків, котрі вирощували на грунті MS з 50–200 мг/л
миш’яку, розвивались нормальні рослини. Отримані in
vitro рослини були стійкими до миш’яку в концентрації
300 мг/кг грунту та накопичували миш’як до 3232 мг/кг
сухої надземної біомаси, що означає покращання рос�
тових характеристик, формування біомаси та накопи�
чення миш’яку в порівнянні з рослинами, вирощени�
ми на полі.
REFERENCES
1. Tamaki S., Frankenberger W.T. 1992. Environmental
biochemistry of Arsenic. Reviews of Environmental
Contamination and Toxicology 124:79–110.
2. U.S. EPA, 2000a. Environmental Protection Agency,
National Primary Drinking Water Regulations; Arsenic
and Clarifications to Compliance and New Source
Contaminants Monitoring; Proposed Rule. Federal
Register, Part II, 40 CFR Parts 141 and 142, Vol. 65
(121): 38888–38983.
ІSSN 0564–3783. Цитология и генетика. 2008. № 5 29
Characterizations of an ecotype of brake'fern, Pteris vittata, for arsenic tolerance and accumulation
3. U.S. EPA, 2002. Proven Alternatives For Above
Ground Treatment Of Arsenic In Ground Water.
EPA–542�S�02–002, June, 2002.
4. Bech J., Poschenrieder C., Llugany M., Barcelo J., Tume
P., Tobias F.J., Barranzuela J.L., Vasques E.R. 1997.
Arsenic and heavy metal contamination of soil and veg�
etation around a copper mine in northern Peru.
Science of Total Environment 203:83–91.
5. Nriagu J.O. 1994. Arsenic in the Environment Part I:
Cycling and Characterization, John Wiley and Sons,
Canada.
6. Hazardous Waste Consultant, 2002. Technologies for
treating arsenic in groundwater. Hazardous Waste
Consultant 20: 114 –119.
7. Welch A.H., Westjohn D.B., Helsel D.R., Wanty R.B. 2000.
Arsenic in ground water of the United States: occurrence
and geochemistry. Ground Water 38: 589–604.
8. Meharg A.A., Hartley�Whitaker J. 2002. Arsenic uptake
and metabolism in arsenic resistant and non�resistant
plant species. New Phytologist 154:29–43.
9. National Research Council, 1999. National Research
Council. Arsenic in Drinking Water. National Academy
Press: Washington, DC.
10. Jiang Q.Q., Singh B.R. 1994. Effect of different forms
and sources of arsenic on crop yield and arsenic con�
centration. Water Air Soil Pollution 74:321–343.
11. Christen K. 2001. The arsenic threat worsens.
Environmental Science and Technology 35: 286A�291A.
12. Brooks R.R. 1998. Plants that Hyper Accumulate
Heavy Metals. Their role in Phytoremediation,
Microbiology, Archaeology, Mineral Exploration and
Phytomining. CAB International.
13. Raskin I., Kumar N.P.B.A., Dushenkov S., Salt D.E.
1994. Bioconcentration of heavy metals by plants.
Current Opinion in Biotechnology 5:285–290.
14. Baker A.J.M. 1987. Metal tolerance. New Phytologist
106:93–111.
15. Baker A.J.M., Mcgrath S.P, Reeves R.D, Smith J.A.C.
2000. Metal hyper accumulator plants: a review of the
ecology and physiology of a biological resource for phy�
toremediation of metal polluted soils. In: Terry, N.,
Banuelos, G. (Eds.), Phytoremediation of Contaminated
Soil and Water. Lewis, Boca Raton, pp.85–107.
16. U.S. EPA, 2000b. US EPA, Office of Research and
Development. Introduction to Phytoremediation.
EPA/600/R�99/107. 2000.
17. Raskin I, Ensley B.D. 2000. Phytoremediation of Toxic
Metals, John Wiley and Sons, Inc., New York
18. Dahmani�Muller H., Van Oort F., Gelie B., Balabane M.
2000. Strategies of heavy metal uptake by three plant
species growing near a metal smelter. Environmental
Pollution, 109: 231–238.
19. Reeves R.D., Schwarz C., Morel J.L., Edmondson J.
2001. Distribution and metal�accumulating behavior of
Thlaspi caerulescens and associated metallophytes in
France. International J Phytoremediation 3:145–172.
20. Licht L.A, McCutcheon S.C., Wolfe N.L., Carreira L.H.
1995. Phytoremediation of organic and nutrient con�
taminants. Environmental Science and Technology
29:318–323.
21. Porter E.K., Peterson P.J. 1975. Arsenic accumulation
by plants on mine waste (United Kingdom). The
Science of The Total Environment 4: 365–371.
22. Macnair M.R, Cumbes Q. 1987. Evidence that arsenic
tolerance in Holcus lanatus L. is caused by an altered
phosphate uptake system. New Phytologist
107:387–394.
23. Meharg A.A., Macnair M.R. 1991b. Uptake, accumula�
tion and translocation of arsenate in arsenate�tolerant
and non�tolerant Holcus lanatus L. New Phytologist
117: 225–231.
24. Meharg A.A., Macnair M.R. 1991a. The mechanisms of
arsenate tolerance in Deschampsia cespitosa (L.) Beauv.
and Agrostis capillaris L. New Phytologist 119: 291–297.
25. Paliouris G., Hutchinson T.C. 1991. Arsenic, cobalt and
nickel tolerances in two populations of Silene vulgaris
(Moench) Garcke from Ontario, Canada. New
Phytologist 117: 449–459.
26. Sneller F.E.C., Van Heerwaarden L.M., Kraaijeveld�Smit
F.J.L., Ten Bookum W.M., Koevoets P.L.M., Schat H.,
Verkleij J.A.C. 1999. Toxicity of arsenate in Silene vul�
garis, accumulation and degradation of arsenate�
induced phytochelatins. New Phytologist 144: 223–232.
27. Sharples J.M., Meharg A.A., Chambers S.M., Cairney
J.W.G. 2000. Symbiotic solution to arsenic contamina�
tion. Nature 404: 951–952.
28. Bleeker P.M., Schat H., Vooijs R., Verkleij J.A.C., Ernst
W.H.O. 2003. Mechanisms of arsenate tolerance in
Cytisus striatus. New Phytologist 157: 33–38.
29. Pickering I.J., Prince R.C., George M.J., Smith R.D.
George G.N., Salt D.E. 2000. Reduction and coordina�
tion of arsenic in Indian mustard. Plant Physiology
122:1171–1177.
30. Schmöger M.E.V., Oven M., Grill E. 2000. Detoxifica�
tion of arsenic by phytochelatins in plants. Plant
Physiology 122:793–801.
31. Koch I., Wang L., Ollson C., Cullen W.R., Reimer K.J.
2000. The predominance of inorganic arsenic species in
plants from Yellowknife, Northwest Territories,
Canada. Environ Science and Technology 34: 22–26.
32. Meharg A.A. 2003. Variation in arsenic accumulation�
hyperaccumulation in ferns and their allies. New
Phytologist 157: 25–31.
33. Matschullat J. 2000. Arsenic in the geosphere – a
review. Science of Total Environment 249: 297–312.
34. Carbonell�Barrachina A.A., Burlo F., Burgos�
Hernanadez A., Lopez E., Mataix J. 1997. The influence
of arsenic concentration on arsenic accumulation in
tomato and bean plants. Scientia Horticulturae.
71:167–176.
ISSN 0564–3783. Цитология и генетика. 2008. № 530
B.K. Sarangi, T. Chakrabarti
35. Kabata�Pendias A., Pendias H. 1991. Arsenic. In: Trace
Elements In Soils And Plants. CRC Press, Boca Raton,
Florida, pp 203–209.
36. Ma L.Q., Komar K.M. Tu C., Zhang W., Cai Y.,
Kennelly E.D. 2001. A fern that hyperaccumulate
arsenic. Nature 409:579.
37. Baker A.J.M., McGrath S.P., Sidoli C.M.D., Reeves R.D.
1994. The possibility of in�situ heavy metal decontam�
ination of polluted soils using crops of metal accumu�
lating plants. Resource Conservation Recycling 11:
41–49.
38. Tu C., Ma L.Q., Bondada B. 2002. Arsenic accumula�
tion in the hyper accumulator Chinese Brake and its
utilization potential for phytoremediation. Journal of
Environmental Quality 31:1671–1675.
39. Tu C., Ma L.Q. 2002. Effects of arsenic concentrations
and forms on arsenic uptake by the hyper accumulator
Ladder Brake. Journal of Environmental Quality
31:641–647.
40. Zhang W., Cai Y., Tu C., Ma L.Q. 2002. Arsenic speci�
ation and distribution in an arsenic hyperaccumulating
plant. The Science of Total Environment 300:167–177.
41. Visoottiviseth P., Francesconi K., Sridokchan W. 2002.
The potential of Thai indigenous plant species for the
phytoremediation of arsenic contaminated land.
Environmental Pollution 118: 453–461.
42. Francesconi K., Visoottiviseth P., Sridokchan W.,
Goessler W. 2002. Arsenic species in an arsenic hyper�
accumulating fern, Pityrogramma calomelanos: a
potential phytoremediator of arsenic�contaminated
soils. Science of Total Envronment 284:27–35.
43. Gumaelius L., Lahner B., Salt E.D., Banks J.A. 2004.
Arsenic Hyperaccumulation in Gametophytes of Pteris
vittata. A new model system for analysis of arsenic
hyperaccumulation. Plant Physiology 136: 3198–3208.
44. Murashige T., Skoog F. 1962. A revised medium for
rapid growth and bioassays with tobacco tissue cultures.
Physiologia Plantarum 15:473–497.
45. Hoagland D.R., Arnon D.I. 1938. The water culture
method for growing plants without soil. California
Agriculture Experimental Station Circular. 347:461
46. Knudson L. 1946. A nutrient solution for the germina�
tion of orchid seed. Bulletin of American Orchid
Society 15: 214–217.
47. Dushenko W.T., Bright D.A., Reimer K.J. 1995. Arsenic
bioaccumulation and toxicity in aquatic macrophytes
to gold�mine effluent: relationship with environmental
partitioning, metal uptake and nutrients. Aquatic
Botany 50:141–158.
48. Salt D.E., Smith R.D., Raskin I. 1998. Phytoremedi�
ation. Annual Review of Plant Physiology and Plant
Molecular Biology 49: 643–668.
49. Zenk M.H. 1996. Heavy metal detoxification in higher
plants – a review. Gene 179: 21–30.
50. Hartley�Whitaker J., Ainsworth G., Vooijs R., Ten Bookum
W.M., Schat H., Meharg A.A. 2001. Phytochelatins are
involved in differential arsenate tolerance in Holcus
lanatus L. Plant Physiology 126: 299–306.
Received 23.08.07
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Characterizations of an ecotype of brake'fern, Pteris vittata, for arsenic tolerance and accumulation
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