Shaik G Mastan - Academia.edu (original) (raw)

Papers by Shaik G Mastan

Jatropha, Challenges for a New Energy Crop, 2019

Interminably increasing petroleum rates and exhaustion of fossil reserves have ignited a global s... more Interminably increasing petroleum rates and exhaustion of fossil reserves have ignited a global search for substitutes to renewable fuel sources. Many oil-generating plants, crops and trees have been considered for biofuel; among these Jatropha curcas is regarded as one of the most promising oilseed plants as its seeds contain oil content up to 35%. Because fossil oil consumption is increasing day-by-day, there is an urgent need to enhance the oil content. Transgenic technology is one of the advanced techniques that have been applied to enhance oil content and modify the composition of fatty acids in seed oils. Increasing seed oil content can be done by modifying the enzyme’s level expression in the triacylglycerol biosynthetic pathway. In this chapter, an effort is made to highlight the potential of transgenic technology towards the enhancement of the oil content and in altering the candidate gene expression for biosynthesis of triacylglycerol.

Gene, Jan 20, 2018

Begomoviruses belong to the family Geminiviridae are associated with several disease symptoms, su... more Begomoviruses belong to the family Geminiviridae are associated with several disease symptoms, such as mosaic and leaf curling in Jatropha curcas. The molecular characterization of these viral strains will help in developing management strategies to control the disease. In this study, J. curcas that was infected with begomovirus and showed acute leaf curling symptoms were identified. DNA-A segment from pathogenic viral strain was isolated and sequenced. The sequenced genome was assembled and characterized in detail. The full-length DNA-A sequence was covered by primer walking. The genome sequence showed the general organization of DNA-A from begomovirus by the distribution of ORFs in both viral and anti-viral strands. The genome size ranged from 2844 bp-2852 bp. Three strains with minor nucleotide variations were identified, and a phylogenetic analysis was performed by comparing the DNA-A segments from other reported begomovirus isolates. The maximum sequence similarity was observed...

Plant Gene, 2016

Jatropha curcas L. (Euphorbiaceae) has gained considerable importance as a renewable source of en... more Jatropha curcas L. (Euphorbiaceae) has gained considerable importance as a renewable source of energy. Various molecular markers were employed to explore the genetic diversity in J. curcas germplasm. In present study, methylation-sensitive AFLP (MS-AFLP) markers were evaluated to analyze the divergence among five elite germplasm (CSMCRI-1 to 5) of J. curcas and subsequently the results were compared with that of AFLP markers to assess the potential utility of MS-AFLP markers for phylogenetic characterization. Study compared the genetic (obtained through AFLP marker) polymorphism with that of epigenetic (obtained through MS-AFLP marker) polymorphism. Among the germplasms studied, 07 selective combinations of MS-AFLP primers produced 40 epigenetic (methylation) polymorphic markers, while 17 AFLP primers produced 105 genetic polymorphic markers. MS-AFLP analysis showed 44.44% epigenetic polymorphism while AFLP showed 15.20% genetic polymorphism. Epigenetic and genetic polymorphism among the germplasms ranged from 12.50% to 66.67% and 2.27% to 48.57% respectively. Highest mean epigenetic polymorphism was recorded in CSMCRI-3, while highest mean genetic polymorphsim was recorded in CSMCRI-1. Results showed that the genetic and epigenetic structures of J. curcas were not same and epigenetic variations are greater than the genetic variations. Differences in DNA methylation and polymorphism among the studied genotypes under prevailing environment indicate evolution of genetic and epigenetic components in different directions. The study paves idea that genotypes exhibiting less genetic divergence might possess high epigenetic divergence and this along with genetic background could be a base for selection of elite cultivar adapted to particular geo-climatic conditions for further breeding program.

Plant Gene, 2016

DNA methylation plays an important role in gene regulation during growth, development and differe... more DNA methylation plays an important role in gene regulation during growth, development and different stresses. We have studied the effect of transgene induced DNA methylation pattern in SbDREB2A overexpressed tobacco transgenic lines. DREB (Dehydration-responsive element binding) transcription factor (TF) plays a vital role in providing stress tolerance in ABA-independent manner by regulating downstream genes in plants. An increase in percent methylation was observed in the transgenic lines as compared to wild type (WT) and plants transformed with vector alone (VA). Among the three transgenic lines, S-201 showed maximum methylation and methylation polymorphism, whereas, the demethylation pattern was observed almost similar for all the transgenic lines. This indicates that DREB gene might be involved during methylation events in transgenic plants by regulating certain regions of the plant genome that plays important role during stress conditions.

International Journal of Environmental Studies, 2015

ABSTRACT The cultivation in wastelands of Jatropha curcas as a biofuel crop avoids the alleged fo... more ABSTRACT The cultivation in wastelands of Jatropha curcas as a biofuel crop avoids the alleged food vs. fuel dilemma. Converting wastelands into Jatropha plantation will produce changes in microbial composition which in turn may have a profound effect on biochemical activities and physico-chemical properties of soil. These changes may be permanent and the soil may attain a different equilibrium through this land use change. The objective of this study was to compare the microbial diversity of native undisturbed soil of barren rocky wasteland and soil drawn from 24 months old J. curcas plantation. Cultivation of Jatropha employing suitable agronomic practices improved soil organic carbon, available P and K. Total soil DNA extracts were used as template DNA to amplify 16S and 18S rDNA gene fragments that were further sequenced and taxonomically assigned by comparisons with gene bank resources. Diversity indices showed that the microbial diversity was higher and more evenly distributed in native soil than in Jatropha planted soil. A prominent diversity shift from Ascomycota in the native soil to Basidiomycota and Chytridiomycota in the Jatropha bulk soils was observed. Interestingly, the conversion from barren land to Jatropha cultivation with recommended agricultural practices also brought about a marked decrease in population of several fungal pathogens. There was a striking increase in members of Proteobacteria (1.7-fold) as well as of Bacteroidetes in Jatropha planted soil as compared to native soil. In contrast, there was a decrease in the Acidobacteria and Chloroflexi community in Jatropha planted soil. It would be useful to follow the microbial pattern over the long term and to study the evolution of the Jatropha soil ecosystem on wastelands.

Molecular Biology Reports, 2014

We investigated DNA methylation and polymorphism in the methylated DNA using AFLP based methylati... more We investigated DNA methylation and polymorphism in the methylated DNA using AFLP based methylation-sensitive amplification polymorphism (MS-AFLP) markers in ecotypes of Jatropha curcas L. growing in similar and different geo-ecological conditions. Three ecotypes growing in different geo-ecological conditions with environmental heterogeneity (Group-1) and five ecotypes growing in similar environmental conditions (Group-2) were assessed. In ecotypes growing in group-1, 44.32 % DNA was methylated and of which 93.59 % DNA was polymorphic. While in group-2, 32.27 % DNA was methylated, of which 51.64 % DNA was polymorphic. In site 1 and site 2 of group-1, overall methylation was 18.94 and 22.44 % respectively with difference of 3.5 %, while overall polymorphism was 41.14 and 39.23 % with a difference of 1.91 %. In site 1 and site 2 of group-2, overall methylation was 24.68 and 24.18 % respectively with difference of 0.5 %, while overall polymorphism was 12.19 and 12.65 % with a difference of 0.46 %. The difference of methylation percentage and percentage of methylation polymorphism throughout the genome of J. curcas at site 1 and 2 of group-1 is higher than that of J. curcas at site 1 and 2 of group-2. These results correlated the physico-chemical properties of soil at these sites. The variations of physico-chemical properties of soil at Chorwadla (site 1 in group-1 and site 2 in group-2) compared to the soil at Brahmapur (site 2 in group-1) is higher than that of soil at Neswad (site 1 in group-2). The study suggests that these homologous nucleotide sequences probably play important role in ecotype adaptation to environmental heterogeneity by creating epiallelic variations hence in evolution of ecotypes/clines or forms of species showing phenotypic/genotypic differences in different geographical areas.

Biosciences Biotechnology Research Asia, 2011

In the present study, a total of four species of biodegrading bacteria namely Bacillus subtilis, ... more In the present study, a total of four species of biodegrading bacteria namely Bacillus subtilis, Eschericia coli, Staphylococcus aereus and Pseudomonas putida were isolated from soil samples collected from the sites where lead and mercury were discharged. The effect of various physicochemical parameters on growth of these bacteria was also studied.

Biomedical & Pharmacology Journal, 2011

In the present study, a total of four species of biodegrading bacteria namely Bacillus subtilis, ... more In the present study, a total of four species of biodegrading bacteria namely Bacillus subtilis, Eschericia coli, Staphylococcus aereus and Pseudomonas putida were isolated from soil samples collected from the sites where lead and mercury were discharged. 16s RNA is amplified using Polymerase Chain Reaction and the amplified product was confirmed by agarose gel electrophoresis and it has been subjected to sequencing and the sequence obtained was compared with the sequence obtained from the nucleotide database of NCBI

Plant Growth Regulation, 2014

In present study, methylation-sensitive AFLP (MSAP) markers were employed to assess DNA methylati... more In present study, methylation-sensitive AFLP (MSAP) markers were employed to assess DNA methylation, degree of alterations in DNA methylation and methylation polymorphism in plant tissues growing in vivo and in vitro. The leaf tissues of six plants growing in vivo and in vitro were subjected to MSAP profiling. A total of 717 MSAP markers in Salvadora persica, 801 in Commiphora wightii, 874 in male (M) and 845 in female (F) genotype of Simmondsia chinensis, 719 in Jatropha curcas and 880 in Withania coagulans were obtained with seventeen MSAP primer combinations. Percentage methylation in genome obtained was higher in in vivo-grown tissues of S. persica (39.47 %), S. chinensis-M (61.71 %) and W. coagulans (71.59 %); and in in vitro-grown tissues of C. wightii (65.17 %), S. chinensis-F (60.83 %) and J. curcas (68.29 %). The percentage polymorphism in methylated DNA obtained was 8.71 % in S. persica, 9.81 % in J. curcas, 10.10 % in S. chinensis-F, 10.26 % in W. coagulans, 10.66 % in S. chinensis-M and 13.98 % in C. wightii. The difference in DNA methylation and polymorphism in genomes reflect the plasticity in genomes of the plants growing under two different environments. Different pattern of DNA methylation of the homologous nucleotide sequences and polymorphism in the methylated DNA in tissues under in vitro and in vivo conditions suggest possibility of involvement of these fragments in the dynamic processes regulating plant growth and development under prevailing growth conditions. Keywords Epigenetic Á In vitro Á In vivo Á Methylation Á MSAP Á Polymorphism Electronic supplementary material The online version of this article (

Gene, 2012

The present study assesses the changes in DNA methylation in leaf and root tissues of Jatropha cu... more The present study assesses the changes in DNA methylation in leaf and root tissues of Jatropha curcas L., induced by salinity stress using methylation sensitive amplification polymorphism (MSAP) markers. Seedlings of 21 days (d) grown under controlled conditions were subjected to 0–100 mM salinity treatment for 24 h (1 d). Immediate changes in DNA methylation and polymorphism in methylated DNA in whole genome of both leaves and roots were assessed using 10 selective combinations of MSAP primers. In root and leaves 70.06% and 57.89% methylation was observed respectively. Similarly 67.22% and 71.21% polymorphism was observed in methylated DNA from root and leaf tissues respectively. Compared with control, the percentage of methylation and methylation polymorphism in roots of plants under different dosages of salinity was found in the order of 50 mM < 25 mM = 100 mM < 75 mM and 75 mM < 25 mM < 50 mM < 100 mM respectively. Similarly percentage of methylation and methylation polymorphism in leaves of plants treated with different levels of salinity was found in order of 75 mM < 25 mM < 50 mM < 100 mM and 50 mM < 25 mM < 100 mM < 75 mM respectively. The MSAP analysis showed that under salt stress homologous nucleotide sequences in genome from control and salt treated plants of J. curcas showed different patterns of methylation; which suggest that these fragments probably play an important role to induce immediate adaptive responses in Jatropha under salinity stress.

Applied Biochemistry and Biotechnology, 2011

Efficient plantlet regeneration with and without intermediate callus phase was achieved for a sel... more Efficient plantlet regeneration with and without intermediate callus phase was achieved for a selected genotype of Aloe vera L. which is sweet in test and used as a vegetable and source of food. Random amplified polymorphic DNA (RAPD) and inter simple sequence repeats (ISSR) marker assays were employed to evaluate genetic stability of plantlets and validate the most reliable method for true-to-type propagation of sweet aloe, among two regeneration systems developed so far. Despite phenotypic similarities in plantlets produced through both regeneration systems, the differences in genomic constituents of plantlets produced through intermediate callus phase using soft base of inflorescence have been effectively distinguished by RAPD and ISSR markers. No polymorphism was observed in regenerants produced following direct regeneration of axillary buds, whereas 80% and 73.3% of polymorphism were observed in RAPD and ISSR, respectively, in the regenerants produced indirectly from base of the inflorescence axis via an intermediate callus phase. Overall, 86.6% of variations were observed in the plantlets produced via an intermediate callus phase. The occurrence of genetic polymorphism is associated with choice of explants and method used for plantlet regeneration. This confirms that clonal propagation of sweet aloe using axillary shoot buds can be used for commercial exploitation of the selected genotype where a high degree of fidelity is an essential prerequisite. On the other hand, a high degree of variations were observed in plantlets obtained through indirect regeneration and thus cannot be used for the mass multiplication of the genotype; however, it can be used for crop improvement through induction of somaclonal variations and genetic manipulations.

Applied Biochemistry and Biotechnology, 2013

The present investigation aimed to evaluate the reliability of in vitro propagation methods for e... more The present investigation aimed to evaluate the reliability of in vitro propagation methods for elite genotypes of Jatropha curcas L., that maintain genetic integrity of tissue culture (TC) regenerates among two regeneration systems developed through direct shoot bud regeneration using nodal/apical shoot segments (protocol-A) and in vitro-derived leaves (protocol-B) as explants. Random amplified polymorphic DNA (RAPD), intersimple sequence repeat (ISSR), simple sequence repeat (SSR) molecular markers, and flow cytometery (FCM) were employed to evaluate genetic homogeneity in TC-regenerates at different passages of subcultures. RAPD markers showed genetic homogeneity in fifth-generation TC-regenerates of both protocols. ISSR markers showed genetic stability of leaf regenerates (protocol-B) at 10th generation. FCM analysis of TC-regenerates at 10th generation in protocol-B and at 20th generation in both protocols, showed stability of ploidy level. SSR assessment of TC-regenerates at 20th generation in both protocols confirmed genetic homogeneity. The results confirmed the genetic stability of the TC-regenerates and demonstrated the reliability of the regeneration systems developed so far using explants of two different origins, for large-scale multiplication of elite genotypes of Jatropha.

South African Journal of Botany, 2016

Withania coagulans (Stocks) Dunal (Solanaceae) is a critically endangered medicinal plant known f... more Withania coagulans (Stocks) Dunal (Solanaceae) is a critically endangered medicinal plant known for its multiple medicinal properties. We report here an efficient and high frequency plantlet regeneration system through direct organogenesis in W. coagulans using in vitro-derived leaves as explants. On Murashige and Skoog's (MS) medium containing 4.44 μM 6-benzylaminopurine (BAP) 73.7 ± 4.3% explants responded and produced 11.4 ± 0.9 shoot buds per explant. The regenerated shoot buds were elongated (6.7 ± 0.22 cm) on MS medium with 1.11 μM BAP and 0.57 μM indole-3-acetic acid (IAA). The elongated shoots were rooted both in vitro and ex vitro. The regenerates were acclimatized by slow and gradual exposure to different regimes of temperature and relative humidity. The leaf-regenerates were assessed for genetic stability using RAPD and ISSR molecular markers and confirmed true-to-type. The in vitro regeneration system developed would be useful for genetic restoration program as the overexploitation and reproductive failure forced W. coagulans towards the verge of complete extinction. The regeneration method would also be useful for genetic transformation for genetic improvement and improved phyto-pharming.

Molecular Biotechnology

Jatropha curcas L., a multipurpose shrub, has acquired significant economic importance for its se... more Jatropha curcas L., a multipurpose shrub, has acquired significant economic importance for its seed oil which can be converted to biodiesel an emerging alternative to petro-diesel. In addition to the commercial value, it is also having medicinal and even high nutritional value to use as animal fodder which is limited due to the toxicity. Development of molecular marker will enable to differentiate non-toxic from toxic variety of J. curcas in a mixed population and also for quality control since the toxic components of J. curcas has deleterious effect on animals. In the present study, the efforts were made to generate the specific SCAR marker for toxic and/or non-toxic J. curcas from RAPD markers. Among the markers specific for toxic and non-toxic varieties, four were selected, purified, cloned, sequenced, and designed primers out of which one set of primers NT-JC/SCAR I/OPQ15-F and R could able to discriminate the non-toxic with toxic Jatropha by giving expected 430 bp size amplification in non-toxic variety. Furthermore, novel multiplex PCR was designed using the nrDNA ITS primers to overcome the false negatives. Present work also demonstrates utility of the conserved regions of nrDNA coding genes in ruling out the artifacts in PCR-like false negatives frequently occur in SCAR due to various reasons. The specific SCAR markers generated in the present investigation will help to distinguish non-toxic from toxic varieties of J. curcas or vice versa, and isolated marker along with designed multiplex protocol has applications in quality control for selective cultivation of non-toxic variety and will also assist in breeding and molecular mapping studies.

Molecular Biology Reports, 2010

Jatropha curcas L. belongs to family Euphorbiaceae, native to South America and widely distribute... more Jatropha curcas L. belongs to family Euphorbiaceae, native to South America and widely distributed in South and Central America, attained significant importance for its seed oil which can be converted to biodiesel, a renewable energy source alternative to conventional petro-diesel. Very few attempts were made to understand the extent of genetic diversity that exists in J. curcas. Therefore, the present investigation was undertaken to asses the genetic diversity among 28 diverse germplasm collected from distinct geographical areas in India. The overall percentage of polymorphism (PP) was found to be 50.70 and 60.95 by RAPD and AFLP, respectively. The mean PP was found to be 9.72 and 20.57 by RAPD and AFLP, respectively. The mean genetic similarity was observed to be 0.89 by RAPD and 0.88 by AFLP. Among the germplasm JCI20 found to be the most diverged one. The dendrogram analysis of RAPD and AFLP data showed good congruence, but better resolution and more polymorphism was observed with AFLP. When the dendrogram of RAPD was compared with AFLP dendrogram, the major clustering pattern was found to be similar; however, changes in minor grouping were observed. In both RAPD and AFLP analysis clustering of germplasm did not show any correlation with the geographical area of collection. Low genetic diversity observed in J. curcas and the clustering pattern indicates that the distribution of species might have happened through anthropogenic activity and warrants the need for widening the genetic base. The present study will provide pavement for further intra-population studies on narrow geographical areas, to understand the population genetic structure, phylogeography, molecular ecological studies. The marker information and the characterized germplasm help in further improvement of the species through marker assisted breeding programs.

Molecular Biology Reports, 2009

Jatropha curcas L., a multipurpose shrub has acquired significant economic importance for its see... more Jatropha curcas L., a multipurpose shrub has acquired significant economic importance for its seed oil which can be converted to biodiesel, is emerging as an alternative to petro-diesel. The deoiled seed cake remains after oil extraction is toxic and cannot be used as a feed despite having best nutritional contents. No quantitative and qualitative differences were observed between toxic and non-toxic varieties of J. curcas except for phorbol esters content. Development of molecular marker will enable to differentiate non-toxic from toxic variety in a mixed population and also help in improvement of the species through marker assisted breeding programs. The present investigation was undertaken to characterize the toxic and non-toxic varieties at molecular level and to develop PCR based molecular markers for distinguishing non-toxic from toxic or vice versa. The polymorphic markers were successfully identified specific to non-toxic and toxic variety using RAPD and AFLP techniques. Totally 371 RAPD, 1,442 AFLP markers were analyzed and 56 (15.09%) RAPD, 238 (16.49%) AFLP markers were found specific to either of the varieties. Genetic similarity between non-toxic and toxic verity was found to be 0.92 by RAPD and 0.90 by AFLP fingerprinting. In the present study out of 12 microsatellite markers analyzed, seven markers were found polymorphic. Among these seven, jcms21 showed homozygous allele in the toxic variety. The study demonstrated that both RAPD and AFLP techniques were equally competitive in identifying polymorphic markers and differentiating both the varieties of J. curcas. Polymorphism of SSR markers prevailed between the varieties of J. curcas. These RAPD and AFLP identified markers will help in selective cultivation of specific variety and along with SSRs these markers can be exploited for further improvement of the species through breeding and Marker Assisted Selection (MAS).

Molecular Biology Reports, 2010

Jatropha curcas L. belongs to family Euphorbiaceae, native to South America attained significant ... more Jatropha curcas L. belongs to family Euphorbiaceae, native to South America attained significant importance for its seed oil which can be converted to biodiesel, a renewable energy source alternative to conventional petrodiesel. Very few attempts were made to isolate novel microsatellite markers and assessment of the extent of genetic equilibrium and diversity that exists in J. curcas. Therefore, the present investigation was undertaken to isolate the novel microsatellites and access genetic equilibrium, diversity that exists among 44 diverse germplasm collected from distinct geographical areas in India using isolated microsatellites. The overall efficiency of the enrichment of microsatellite by dual probe in the present study found to be 54% and among the sequences obtained the percentage of sequences having suitable flanking regions for the primer designing was found to be 89.58%. The mean co-efficient of genetic similarity (CGS) was found to be 0.97. The overall diversity obtained by microsatellites was found to be low in comparison with the diversity reported by multilocus markers systems observed in earlier studies; however, the good allele polymorphism was observed. The overall dendrogram of microsatellite analysis resulted in random clustering of germplasm and not in accordance to geographical area of collection. The present study, diversity analysis using microsatellite markers concludes the low genetic diversity and genetic disequlibrium of J. curcas in India and will provide pavement for further intra-population studies on narrow geographical areas to understand the population genetic structure, phylogeography and molecular ecological studies. The germplasm characterized, and the microsatellite markers isolated and characterized in the present study can be employed efficiently in breeding programs for genetic improvement of the species through marker assisted selection and QTL analysis, for further genetic resource management and help in making the J. curcas as potential crop with superior agronomical traits.

Molecular Biology Reports, 2011

The present investigation was undertaken with an aim to check the ability of cross species amplif... more The present investigation was undertaken with an aim to check the ability of cross species amplification of microsatellite markers isolated from Jatropha curcas—a renewable source of biodiesel to deduce the generic relationship with its six sister taxa (J. glandulifera, J. gossypifolia, J. integerrima, J. multifida, J. podagrica, and J. tanjorensis). Out of the 49 markers checked 31 markers showed cross species amplification in all the species studied. JCDS-30, JCDS-69, JCDS-26, JCMS-13 and JCMS-21 amplified in J. curcas. However, these markers did not show any cross species amplification. Overall percentage of polymorphism (PP) among the species studied was 38% and the mean genetic similarity (GS) was found to be 0.86. The highest PP (24) and least GS (0.76) was found between J. curcas/J. podagrica and J. curcas/J. multifida and least PP (4.44) and highest GS (0.96) was found between J. integerrima/J. tanjorensis. Dendrogram analysis showed good congruence to RAPD and AFLP than nrDNA ITS data reported earlier. The characterized microsatellites will pave way for intraspecies molecular characterization which can be further utilized in species differentiation, molecular identification, characterization of interspecific hybrids, exploitation of genetic resource management and genetic improvement of the species through marker assisted breeding for economically important traits.

Jatropha, Challenges for a New Energy Crop, 2019

Interminably increasing petroleum rates and exhaustion of fossil reserves have ignited a global s... more Interminably increasing petroleum rates and exhaustion of fossil reserves have ignited a global search for substitutes to renewable fuel sources. Many oil-generating plants, crops and trees have been considered for biofuel; among these Jatropha curcas is regarded as one of the most promising oilseed plants as its seeds contain oil content up to 35%. Because fossil oil consumption is increasing day-by-day, there is an urgent need to enhance the oil content. Transgenic technology is one of the advanced techniques that have been applied to enhance oil content and modify the composition of fatty acids in seed oils. Increasing seed oil content can be done by modifying the enzyme’s level expression in the triacylglycerol biosynthetic pathway. In this chapter, an effort is made to highlight the potential of transgenic technology towards the enhancement of the oil content and in altering the candidate gene expression for biosynthesis of triacylglycerol.

Gene, Jan 20, 2018

Begomoviruses belong to the family Geminiviridae are associated with several disease symptoms, su... more Begomoviruses belong to the family Geminiviridae are associated with several disease symptoms, such as mosaic and leaf curling in Jatropha curcas. The molecular characterization of these viral strains will help in developing management strategies to control the disease. In this study, J. curcas that was infected with begomovirus and showed acute leaf curling symptoms were identified. DNA-A segment from pathogenic viral strain was isolated and sequenced. The sequenced genome was assembled and characterized in detail. The full-length DNA-A sequence was covered by primer walking. The genome sequence showed the general organization of DNA-A from begomovirus by the distribution of ORFs in both viral and anti-viral strands. The genome size ranged from 2844 bp-2852 bp. Three strains with minor nucleotide variations were identified, and a phylogenetic analysis was performed by comparing the DNA-A segments from other reported begomovirus isolates. The maximum sequence similarity was observed...

Plant Gene, 2016

Jatropha curcas L. (Euphorbiaceae) has gained considerable importance as a renewable source of en... more Jatropha curcas L. (Euphorbiaceae) has gained considerable importance as a renewable source of energy. Various molecular markers were employed to explore the genetic diversity in J. curcas germplasm. In present study, methylation-sensitive AFLP (MS-AFLP) markers were evaluated to analyze the divergence among five elite germplasm (CSMCRI-1 to 5) of J. curcas and subsequently the results were compared with that of AFLP markers to assess the potential utility of MS-AFLP markers for phylogenetic characterization. Study compared the genetic (obtained through AFLP marker) polymorphism with that of epigenetic (obtained through MS-AFLP marker) polymorphism. Among the germplasms studied, 07 selective combinations of MS-AFLP primers produced 40 epigenetic (methylation) polymorphic markers, while 17 AFLP primers produced 105 genetic polymorphic markers. MS-AFLP analysis showed 44.44% epigenetic polymorphism while AFLP showed 15.20% genetic polymorphism. Epigenetic and genetic polymorphism among the germplasms ranged from 12.50% to 66.67% and 2.27% to 48.57% respectively. Highest mean epigenetic polymorphism was recorded in CSMCRI-3, while highest mean genetic polymorphsim was recorded in CSMCRI-1. Results showed that the genetic and epigenetic structures of J. curcas were not same and epigenetic variations are greater than the genetic variations. Differences in DNA methylation and polymorphism among the studied genotypes under prevailing environment indicate evolution of genetic and epigenetic components in different directions. The study paves idea that genotypes exhibiting less genetic divergence might possess high epigenetic divergence and this along with genetic background could be a base for selection of elite cultivar adapted to particular geo-climatic conditions for further breeding program.

Plant Gene, 2016

DNA methylation plays an important role in gene regulation during growth, development and differe... more DNA methylation plays an important role in gene regulation during growth, development and different stresses. We have studied the effect of transgene induced DNA methylation pattern in SbDREB2A overexpressed tobacco transgenic lines. DREB (Dehydration-responsive element binding) transcription factor (TF) plays a vital role in providing stress tolerance in ABA-independent manner by regulating downstream genes in plants. An increase in percent methylation was observed in the transgenic lines as compared to wild type (WT) and plants transformed with vector alone (VA). Among the three transgenic lines, S-201 showed maximum methylation and methylation polymorphism, whereas, the demethylation pattern was observed almost similar for all the transgenic lines. This indicates that DREB gene might be involved during methylation events in transgenic plants by regulating certain regions of the plant genome that plays important role during stress conditions.

International Journal of Environmental Studies, 2015

ABSTRACT The cultivation in wastelands of Jatropha curcas as a biofuel crop avoids the alleged fo... more ABSTRACT The cultivation in wastelands of Jatropha curcas as a biofuel crop avoids the alleged food vs. fuel dilemma. Converting wastelands into Jatropha plantation will produce changes in microbial composition which in turn may have a profound effect on biochemical activities and physico-chemical properties of soil. These changes may be permanent and the soil may attain a different equilibrium through this land use change. The objective of this study was to compare the microbial diversity of native undisturbed soil of barren rocky wasteland and soil drawn from 24 months old J. curcas plantation. Cultivation of Jatropha employing suitable agronomic practices improved soil organic carbon, available P and K. Total soil DNA extracts were used as template DNA to amplify 16S and 18S rDNA gene fragments that were further sequenced and taxonomically assigned by comparisons with gene bank resources. Diversity indices showed that the microbial diversity was higher and more evenly distributed in native soil than in Jatropha planted soil. A prominent diversity shift from Ascomycota in the native soil to Basidiomycota and Chytridiomycota in the Jatropha bulk soils was observed. Interestingly, the conversion from barren land to Jatropha cultivation with recommended agricultural practices also brought about a marked decrease in population of several fungal pathogens. There was a striking increase in members of Proteobacteria (1.7-fold) as well as of Bacteroidetes in Jatropha planted soil as compared to native soil. In contrast, there was a decrease in the Acidobacteria and Chloroflexi community in Jatropha planted soil. It would be useful to follow the microbial pattern over the long term and to study the evolution of the Jatropha soil ecosystem on wastelands.

Molecular Biology Reports, 2014

We investigated DNA methylation and polymorphism in the methylated DNA using AFLP based methylati... more We investigated DNA methylation and polymorphism in the methylated DNA using AFLP based methylation-sensitive amplification polymorphism (MS-AFLP) markers in ecotypes of Jatropha curcas L. growing in similar and different geo-ecological conditions. Three ecotypes growing in different geo-ecological conditions with environmental heterogeneity (Group-1) and five ecotypes growing in similar environmental conditions (Group-2) were assessed. In ecotypes growing in group-1, 44.32 % DNA was methylated and of which 93.59 % DNA was polymorphic. While in group-2, 32.27 % DNA was methylated, of which 51.64 % DNA was polymorphic. In site 1 and site 2 of group-1, overall methylation was 18.94 and 22.44 % respectively with difference of 3.5 %, while overall polymorphism was 41.14 and 39.23 % with a difference of 1.91 %. In site 1 and site 2 of group-2, overall methylation was 24.68 and 24.18 % respectively with difference of 0.5 %, while overall polymorphism was 12.19 and 12.65 % with a difference of 0.46 %. The difference of methylation percentage and percentage of methylation polymorphism throughout the genome of J. curcas at site 1 and 2 of group-1 is higher than that of J. curcas at site 1 and 2 of group-2. These results correlated the physico-chemical properties of soil at these sites. The variations of physico-chemical properties of soil at Chorwadla (site 1 in group-1 and site 2 in group-2) compared to the soil at Brahmapur (site 2 in group-1) is higher than that of soil at Neswad (site 1 in group-2). The study suggests that these homologous nucleotide sequences probably play important role in ecotype adaptation to environmental heterogeneity by creating epiallelic variations hence in evolution of ecotypes/clines or forms of species showing phenotypic/genotypic differences in different geographical areas.

Biosciences Biotechnology Research Asia, 2011

In the present study, a total of four species of biodegrading bacteria namely Bacillus subtilis, ... more In the present study, a total of four species of biodegrading bacteria namely Bacillus subtilis, Eschericia coli, Staphylococcus aereus and Pseudomonas putida were isolated from soil samples collected from the sites where lead and mercury were discharged. The effect of various physicochemical parameters on growth of these bacteria was also studied.

Biomedical & Pharmacology Journal, 2011

In the present study, a total of four species of biodegrading bacteria namely Bacillus subtilis, ... more In the present study, a total of four species of biodegrading bacteria namely Bacillus subtilis, Eschericia coli, Staphylococcus aereus and Pseudomonas putida were isolated from soil samples collected from the sites where lead and mercury were discharged. 16s RNA is amplified using Polymerase Chain Reaction and the amplified product was confirmed by agarose gel electrophoresis and it has been subjected to sequencing and the sequence obtained was compared with the sequence obtained from the nucleotide database of NCBI

Plant Growth Regulation, 2014

In present study, methylation-sensitive AFLP (MSAP) markers were employed to assess DNA methylati... more In present study, methylation-sensitive AFLP (MSAP) markers were employed to assess DNA methylation, degree of alterations in DNA methylation and methylation polymorphism in plant tissues growing in vivo and in vitro. The leaf tissues of six plants growing in vivo and in vitro were subjected to MSAP profiling. A total of 717 MSAP markers in Salvadora persica, 801 in Commiphora wightii, 874 in male (M) and 845 in female (F) genotype of Simmondsia chinensis, 719 in Jatropha curcas and 880 in Withania coagulans were obtained with seventeen MSAP primer combinations. Percentage methylation in genome obtained was higher in in vivo-grown tissues of S. persica (39.47 %), S. chinensis-M (61.71 %) and W. coagulans (71.59 %); and in in vitro-grown tissues of C. wightii (65.17 %), S. chinensis-F (60.83 %) and J. curcas (68.29 %). The percentage polymorphism in methylated DNA obtained was 8.71 % in S. persica, 9.81 % in J. curcas, 10.10 % in S. chinensis-F, 10.26 % in W. coagulans, 10.66 % in S. chinensis-M and 13.98 % in C. wightii. The difference in DNA methylation and polymorphism in genomes reflect the plasticity in genomes of the plants growing under two different environments. Different pattern of DNA methylation of the homologous nucleotide sequences and polymorphism in the methylated DNA in tissues under in vitro and in vivo conditions suggest possibility of involvement of these fragments in the dynamic processes regulating plant growth and development under prevailing growth conditions. Keywords Epigenetic Á In vitro Á In vivo Á Methylation Á MSAP Á Polymorphism Electronic supplementary material The online version of this article (

Gene, 2012

The present study assesses the changes in DNA methylation in leaf and root tissues of Jatropha cu... more The present study assesses the changes in DNA methylation in leaf and root tissues of Jatropha curcas L., induced by salinity stress using methylation sensitive amplification polymorphism (MSAP) markers. Seedlings of 21 days (d) grown under controlled conditions were subjected to 0–100 mM salinity treatment for 24 h (1 d). Immediate changes in DNA methylation and polymorphism in methylated DNA in whole genome of both leaves and roots were assessed using 10 selective combinations of MSAP primers. In root and leaves 70.06% and 57.89% methylation was observed respectively. Similarly 67.22% and 71.21% polymorphism was observed in methylated DNA from root and leaf tissues respectively. Compared with control, the percentage of methylation and methylation polymorphism in roots of plants under different dosages of salinity was found in the order of 50 mM < 25 mM = 100 mM < 75 mM and 75 mM < 25 mM < 50 mM < 100 mM respectively. Similarly percentage of methylation and methylation polymorphism in leaves of plants treated with different levels of salinity was found in order of 75 mM < 25 mM < 50 mM < 100 mM and 50 mM < 25 mM < 100 mM < 75 mM respectively. The MSAP analysis showed that under salt stress homologous nucleotide sequences in genome from control and salt treated plants of J. curcas showed different patterns of methylation; which suggest that these fragments probably play an important role to induce immediate adaptive responses in Jatropha under salinity stress.

Applied Biochemistry and Biotechnology, 2011

Efficient plantlet regeneration with and without intermediate callus phase was achieved for a sel... more Efficient plantlet regeneration with and without intermediate callus phase was achieved for a selected genotype of Aloe vera L. which is sweet in test and used as a vegetable and source of food. Random amplified polymorphic DNA (RAPD) and inter simple sequence repeats (ISSR) marker assays were employed to evaluate genetic stability of plantlets and validate the most reliable method for true-to-type propagation of sweet aloe, among two regeneration systems developed so far. Despite phenotypic similarities in plantlets produced through both regeneration systems, the differences in genomic constituents of plantlets produced through intermediate callus phase using soft base of inflorescence have been effectively distinguished by RAPD and ISSR markers. No polymorphism was observed in regenerants produced following direct regeneration of axillary buds, whereas 80% and 73.3% of polymorphism were observed in RAPD and ISSR, respectively, in the regenerants produced indirectly from base of the inflorescence axis via an intermediate callus phase. Overall, 86.6% of variations were observed in the plantlets produced via an intermediate callus phase. The occurrence of genetic polymorphism is associated with choice of explants and method used for plantlet regeneration. This confirms that clonal propagation of sweet aloe using axillary shoot buds can be used for commercial exploitation of the selected genotype where a high degree of fidelity is an essential prerequisite. On the other hand, a high degree of variations were observed in plantlets obtained through indirect regeneration and thus cannot be used for the mass multiplication of the genotype; however, it can be used for crop improvement through induction of somaclonal variations and genetic manipulations.

Applied Biochemistry and Biotechnology, 2013

The present investigation aimed to evaluate the reliability of in vitro propagation methods for e... more The present investigation aimed to evaluate the reliability of in vitro propagation methods for elite genotypes of Jatropha curcas L., that maintain genetic integrity of tissue culture (TC) regenerates among two regeneration systems developed through direct shoot bud regeneration using nodal/apical shoot segments (protocol-A) and in vitro-derived leaves (protocol-B) as explants. Random amplified polymorphic DNA (RAPD), intersimple sequence repeat (ISSR), simple sequence repeat (SSR) molecular markers, and flow cytometery (FCM) were employed to evaluate genetic homogeneity in TC-regenerates at different passages of subcultures. RAPD markers showed genetic homogeneity in fifth-generation TC-regenerates of both protocols. ISSR markers showed genetic stability of leaf regenerates (protocol-B) at 10th generation. FCM analysis of TC-regenerates at 10th generation in protocol-B and at 20th generation in both protocols, showed stability of ploidy level. SSR assessment of TC-regenerates at 20th generation in both protocols confirmed genetic homogeneity. The results confirmed the genetic stability of the TC-regenerates and demonstrated the reliability of the regeneration systems developed so far using explants of two different origins, for large-scale multiplication of elite genotypes of Jatropha.

South African Journal of Botany, 2016

Withania coagulans (Stocks) Dunal (Solanaceae) is a critically endangered medicinal plant known f... more Withania coagulans (Stocks) Dunal (Solanaceae) is a critically endangered medicinal plant known for its multiple medicinal properties. We report here an efficient and high frequency plantlet regeneration system through direct organogenesis in W. coagulans using in vitro-derived leaves as explants. On Murashige and Skoog's (MS) medium containing 4.44 μM 6-benzylaminopurine (BAP) 73.7 ± 4.3% explants responded and produced 11.4 ± 0.9 shoot buds per explant. The regenerated shoot buds were elongated (6.7 ± 0.22 cm) on MS medium with 1.11 μM BAP and 0.57 μM indole-3-acetic acid (IAA). The elongated shoots were rooted both in vitro and ex vitro. The regenerates were acclimatized by slow and gradual exposure to different regimes of temperature and relative humidity. The leaf-regenerates were assessed for genetic stability using RAPD and ISSR molecular markers and confirmed true-to-type. The in vitro regeneration system developed would be useful for genetic restoration program as the overexploitation and reproductive failure forced W. coagulans towards the verge of complete extinction. The regeneration method would also be useful for genetic transformation for genetic improvement and improved phyto-pharming.

Molecular Biotechnology

Jatropha curcas L., a multipurpose shrub, has acquired significant economic importance for its se... more Jatropha curcas L., a multipurpose shrub, has acquired significant economic importance for its seed oil which can be converted to biodiesel an emerging alternative to petro-diesel. In addition to the commercial value, it is also having medicinal and even high nutritional value to use as animal fodder which is limited due to the toxicity. Development of molecular marker will enable to differentiate non-toxic from toxic variety of J. curcas in a mixed population and also for quality control since the toxic components of J. curcas has deleterious effect on animals. In the present study, the efforts were made to generate the specific SCAR marker for toxic and/or non-toxic J. curcas from RAPD markers. Among the markers specific for toxic and non-toxic varieties, four were selected, purified, cloned, sequenced, and designed primers out of which one set of primers NT-JC/SCAR I/OPQ15-F and R could able to discriminate the non-toxic with toxic Jatropha by giving expected 430 bp size amplification in non-toxic variety. Furthermore, novel multiplex PCR was designed using the nrDNA ITS primers to overcome the false negatives. Present work also demonstrates utility of the conserved regions of nrDNA coding genes in ruling out the artifacts in PCR-like false negatives frequently occur in SCAR due to various reasons. The specific SCAR markers generated in the present investigation will help to distinguish non-toxic from toxic varieties of J. curcas or vice versa, and isolated marker along with designed multiplex protocol has applications in quality control for selective cultivation of non-toxic variety and will also assist in breeding and molecular mapping studies.

Molecular Biology Reports, 2010

Jatropha curcas L. belongs to family Euphorbiaceae, native to South America and widely distribute... more Jatropha curcas L. belongs to family Euphorbiaceae, native to South America and widely distributed in South and Central America, attained significant importance for its seed oil which can be converted to biodiesel, a renewable energy source alternative to conventional petro-diesel. Very few attempts were made to understand the extent of genetic diversity that exists in J. curcas. Therefore, the present investigation was undertaken to asses the genetic diversity among 28 diverse germplasm collected from distinct geographical areas in India. The overall percentage of polymorphism (PP) was found to be 50.70 and 60.95 by RAPD and AFLP, respectively. The mean PP was found to be 9.72 and 20.57 by RAPD and AFLP, respectively. The mean genetic similarity was observed to be 0.89 by RAPD and 0.88 by AFLP. Among the germplasm JCI20 found to be the most diverged one. The dendrogram analysis of RAPD and AFLP data showed good congruence, but better resolution and more polymorphism was observed with AFLP. When the dendrogram of RAPD was compared with AFLP dendrogram, the major clustering pattern was found to be similar; however, changes in minor grouping were observed. In both RAPD and AFLP analysis clustering of germplasm did not show any correlation with the geographical area of collection. Low genetic diversity observed in J. curcas and the clustering pattern indicates that the distribution of species might have happened through anthropogenic activity and warrants the need for widening the genetic base. The present study will provide pavement for further intra-population studies on narrow geographical areas, to understand the population genetic structure, phylogeography, molecular ecological studies. The marker information and the characterized germplasm help in further improvement of the species through marker assisted breeding programs.

Molecular Biology Reports, 2009

Jatropha curcas L., a multipurpose shrub has acquired significant economic importance for its see... more Jatropha curcas L., a multipurpose shrub has acquired significant economic importance for its seed oil which can be converted to biodiesel, is emerging as an alternative to petro-diesel. The deoiled seed cake remains after oil extraction is toxic and cannot be used as a feed despite having best nutritional contents. No quantitative and qualitative differences were observed between toxic and non-toxic varieties of J. curcas except for phorbol esters content. Development of molecular marker will enable to differentiate non-toxic from toxic variety in a mixed population and also help in improvement of the species through marker assisted breeding programs. The present investigation was undertaken to characterize the toxic and non-toxic varieties at molecular level and to develop PCR based molecular markers for distinguishing non-toxic from toxic or vice versa. The polymorphic markers were successfully identified specific to non-toxic and toxic variety using RAPD and AFLP techniques. Totally 371 RAPD, 1,442 AFLP markers were analyzed and 56 (15.09%) RAPD, 238 (16.49%) AFLP markers were found specific to either of the varieties. Genetic similarity between non-toxic and toxic verity was found to be 0.92 by RAPD and 0.90 by AFLP fingerprinting. In the present study out of 12 microsatellite markers analyzed, seven markers were found polymorphic. Among these seven, jcms21 showed homozygous allele in the toxic variety. The study demonstrated that both RAPD and AFLP techniques were equally competitive in identifying polymorphic markers and differentiating both the varieties of J. curcas. Polymorphism of SSR markers prevailed between the varieties of J. curcas. These RAPD and AFLP identified markers will help in selective cultivation of specific variety and along with SSRs these markers can be exploited for further improvement of the species through breeding and Marker Assisted Selection (MAS).

Molecular Biology Reports, 2010

Jatropha curcas L. belongs to family Euphorbiaceae, native to South America attained significant ... more Jatropha curcas L. belongs to family Euphorbiaceae, native to South America attained significant importance for its seed oil which can be converted to biodiesel, a renewable energy source alternative to conventional petrodiesel. Very few attempts were made to isolate novel microsatellite markers and assessment of the extent of genetic equilibrium and diversity that exists in J. curcas. Therefore, the present investigation was undertaken to isolate the novel microsatellites and access genetic equilibrium, diversity that exists among 44 diverse germplasm collected from distinct geographical areas in India using isolated microsatellites. The overall efficiency of the enrichment of microsatellite by dual probe in the present study found to be 54% and among the sequences obtained the percentage of sequences having suitable flanking regions for the primer designing was found to be 89.58%. The mean co-efficient of genetic similarity (CGS) was found to be 0.97. The overall diversity obtained by microsatellites was found to be low in comparison with the diversity reported by multilocus markers systems observed in earlier studies; however, the good allele polymorphism was observed. The overall dendrogram of microsatellite analysis resulted in random clustering of germplasm and not in accordance to geographical area of collection. The present study, diversity analysis using microsatellite markers concludes the low genetic diversity and genetic disequlibrium of J. curcas in India and will provide pavement for further intra-population studies on narrow geographical areas to understand the population genetic structure, phylogeography and molecular ecological studies. The germplasm characterized, and the microsatellite markers isolated and characterized in the present study can be employed efficiently in breeding programs for genetic improvement of the species through marker assisted selection and QTL analysis, for further genetic resource management and help in making the J. curcas as potential crop with superior agronomical traits.

Molecular Biology Reports, 2011

The present investigation was undertaken with an aim to check the ability of cross species amplif... more The present investigation was undertaken with an aim to check the ability of cross species amplification of microsatellite markers isolated from Jatropha curcas—a renewable source of biodiesel to deduce the generic relationship with its six sister taxa (J. glandulifera, J. gossypifolia, J. integerrima, J. multifida, J. podagrica, and J. tanjorensis). Out of the 49 markers checked 31 markers showed cross species amplification in all the species studied. JCDS-30, JCDS-69, JCDS-26, JCMS-13 and JCMS-21 amplified in J. curcas. However, these markers did not show any cross species amplification. Overall percentage of polymorphism (PP) among the species studied was 38% and the mean genetic similarity (GS) was found to be 0.86. The highest PP (24) and least GS (0.76) was found between J. curcas/J. podagrica and J. curcas/J. multifida and least PP (4.44) and highest GS (0.96) was found between J. integerrima/J. tanjorensis. Dendrogram analysis showed good congruence to RAPD and AFLP than nrDNA ITS data reported earlier. The characterized microsatellites will pave way for intraspecies molecular characterization which can be further utilized in species differentiation, molecular identification, characterization of interspecific hybrids, exploitation of genetic resource management and genetic improvement of the species through marker assisted breeding for economically important traits.