Browsing by Author "Sumita Jha"

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Chromosomal localization of 45S rDNA, sex-specific C values, and heterochromatin distribution in Coccinia grandis (L.) Voigt
(Springer-Verlag Wien, 2016) Biplab Kumar Bhowmick; Masashi Yamamoto; Sumita Jha
Coccinia grandis is a widely distributed dioecious cucurbit in India, with heteromorphic sex chromosomes and X-Y sex determination mode. The present study aids in the cytogenetic characterization of four native populations of this plant employing distribution patterns of 45S rDNA on chromosomes and guanine-cytosine (GC)-rich heterochromatin in the genome coupled with flow cytometric determination of genome sizes. Existence of four nucleolar chromosomes could be confirmed by the presence of four telomeric 45S rDNA signals in both male and female plants. All four 45S rDNA sites are rich in heterochromatin evident from the co-localization of telomeric chromomycin A (CMA)+ve signals. The size of 45S rDNA signal was found to differ between the homologues of one nucleolar chromosome pair. The distribution of heterochromatin is found to differ among the male and female populations. The average GC-rich heterochromatin content of male and female populations is 23.27 and 29.86 %, respectively. Moreover, the male plants have a genome size of 0.92 pg/2C while the female plants have a size of 0.73 pg/2C, reflecting a huge genomic divergence between the genders. The great variation in genome size is owing to the presence of Y chromosome in the male populations, playing a multifaceted role in sexual divergence in C. grandis. © 2015, Springer-Verlag Wien.
Chromosome morphometric analysis of Indian cultivars of Lens culinaris Medik. using EMA based Giemsa staining method
(Taylor and Francis Ltd., 2017) Timir Baran Jha; Partha Sarathi Saha; Mousumi Adak; Sumita Jha; Partha Roy
The influence of biotic, abiotic and climatic changes on crop production is becoming increasingly evident. Gaps in demand and supply of lentils, one of the few protein-rich crops in India, are gradually increasing. Of seven estimated species of lentil, six are wild and the other, Lens culinaris, is the only species under cultivation, with a large number of cultivars. Chromosome analysis is beneficial to breeders and genome researchers in crop improvement programmes. However, a chromosomal database of this important crop is not available in India. The present paper has described a detailed chromosome analysis of 21 certified Indian cultivars of L. culinaris using enzymatic maceration and air drying (EMA) based Giemsa staining methods for the first time. Uniform chromosomal preparations have resulted in variation in average chromosome size (4.94–9.8 μm), total chromatin length (69.18–137.24 μm), and satellite bearing chromosome number (third, fourth and fifth) among the cultivars. Though our results revealed similar karyotypic formulae and symmetrical karyotype, a scatter diagram of intra-chromosomal asymmetry index (A1) versus inter-chromosomal asymmetry index (A2) groups them into five distinct clusters. Such information may be helpful for conservation of genetic diversity and for future lentil breeding programmes. © 2017 Dipartimento di Biologia Evoluzionistica, Università di Firenze.
Cytogenetics of two Indian varieties of Momordica charantia L. (bittergourd)
(Elsevier B.V., 2018) Ipshita Ghosh; Biplab Kumar Bhowmick; Sumita Jha
Momordica charantia L.commonly known as bitter gourd is a widely cultivated cash crop in India with immense medicinal values. India has been one centre of diversification of two prominent taxonomic varieties viz. long-fruited variety, M. charantia var. charantia (MCC) and small-fruited variety, M. charantia var. muricata (MCM). Momordica species have long been known not to be amenable to conventional cytological techniques. After thorough technical standardization, karyotype variation between the two varieties as well as different populations of the same variety was compared and we primarily structured karyotypes of MCC and MCM populations that conformed to diploid count of 2n = 22 with slightly greater chromosome size in MCM (1.32μm–3.24 μm) than MCC (1.27μm–3.07 μm). Significant differences in genome size between MCC and MCM was also confirmed by flow cytometry for the first time in the two varieties of the species. Pronounced intraspecific dissimilarity in somatic chromosomes was established by EMA-Giemsa method revealing four nucleolar chromosomes in MCC populations and six nucleolar chromosomes in MCM populations. This difference in varieties was substantiated by fluorochrome banding which revealed four and six distal CMA+ve signals in MCC and MCM respectively. Detailed inter- and intra- karyomorphometric evaluation ascertained higher tendency of karyotype asymmetry in MCC. EMA-DAPI technique was applied for meiotic analysis in MCC (n = 11) and MCM (n = 11) populations providing indications of regular meiotic behavior. These results will benefit the classification, the genetic basis of domestication and the genome studies of this widely cultivated cash crop. © 2018 Elsevier B.V.
Effects associated with insertion of cryptogein gene utilizing Ri and Ti plasmids on morphology and secondary metabolites are stable in Bacopa monnieri-transformed plants grown in vitro and ex vitro
(Springer Tokyo, 2015) Pijush Paul; Sayantika Sarkar; Sumita Jha
Bacopa monnieri (Linn.) Wettst. (family Scrophulariaceae), a therapeutically important perennial herb, was transformed to study the stability of the effects associated with the insertion of a gene encoding β-cryptogein, a proteinaceous elicitor, either alone or in addition to the T-DNA genes in long-term in vitro culture as well as after transfer to the greenhouse. Plant lines BmAr-IXcrypt obtained via transformation by LBA 9402-crypt (pRi1855+pBin19++crypt) showed integration and expression of rolA, rolB, rolC, and rolD genes after 4 years of maintenance in vitro. The plant line BmAt-ncrypt obtained via transformation with A. tumefaciens strain LBA4404-crypt (harboring pBin19++crypt) as well as plant line BmAr-IXcrypt showed stable integration and expression of nptII gene and crypt gene even after transfer to greenhouse. The clones of Ri-crypt-transformed plants differed morphologically from Ri-transformed plants in that the typical “hairy root syndrome” was not observed in plants expressing crypt gene in the presence of rol genes. Except plant line BmAr-IXcrypt, none of the other transgenic plant lines showed any alteration in floral morphology and onset of flowering. The crypt-transformed plant lines (BmAr-IXcrypt and BmAt-ncrypt), maintained under long-term culture, showed significantly higher (p ≤ 0.05) amount of bacoside production in vitro (1.66- to 2.05-fold, with respect to non-transformed plants). In addition, accumulation of all the four bacosides was significantly higher (p ≤ 0.05) in crypt-transformed plants as compared to non-transformed plants grown for 1 year in greenhouse. Thus, the crypt-transformed plant lines of B. monnieri may be considered as the potential source for sustainable bioproduction of bacosides. © 2015, Korean Society for Plant Biotechnology and Springer Japan.
Effects of cryptogein gene on growth, phenotype and secondary metabolite accumulation in co-transformed roots and plants of Tylophora indica
(Polish Academy of Sciences, 2017) Amrita Basu; Dipasree Roychowdhury; Raj Kumar Joshi; Sumita Jha
Tylophora indica, an indigenous medicinal plant, was transformed with the cryptogein gene to determine the effect of crypt gene on secondary metabolites in co-transformed roots and plants via Agrobacterium rhizogenes mediated transformation. The Ri crypt co-transformed roots and plants showed expression of crypt gene. Southern hybridization specifies that crypt gene has been transferred and positively integrated into the Ri crypt co-transformed plant. AFLP fingerprinting revealed high degree of genetic similarity among the Ri-transformed and Ri crypt co-transformed cultures. The expression of crypt gene stimulated phenolic compound accumulation in transformed root and plants while tylophorine content was comparable in Ri transformed and Ri crypt co-transformed root lines and plants. The Ri crypt co-transformed root lines showed significantly higher (p ≤ 0.05) phenolics production (caffeic acid, 1.8–2.9-fold; p-coumaric acid, 1.9-fold and ferulic acid, 1.5–2-fold) compared to Ri-transformed root lines. The roots of Ri crypt co-transformed plants showed a significantly (p ≤ 0.05) higher content of caffeic acid (1.19-fold) and ferulic acid (1.53-fold) than Ri-transformed plants. It is suggested that crypt-transformed plants can also be used as a tool to elucidate the biochemical basis of defense responses as phenolics are known to play a role in providing defense barriers to infection by pathogen. © 2016, Franciszek Górski Institute of Plant Physiology, Polish Academy of Sciences, Kraków.
Enhanced trans-resveratrol production in genetically transformed root cultures of Peanut (Arachis hypogaea L.)
(Springer Netherlands, 2016) Mihir Halder; Sumita Jha
In the present study, 30 Ri-transformed root lines of peanut (Arachis hypogaea) cv. JL-24, a popular Indian cultivar, obtained following infection with Agrobacterium rhizogenes strains LBA9402, A4 and R1000 were selected on the basis of growth index and maintained in vitro for 3 years. The root lines showed high degree of branching and rapid, plagiotropic growth on phytohormone free solid N/5 medium but were devoid of root hairs. Trans-resveratrol was isolated by preparative HPLC from Ri-transformed roots and identified by ESI–MS/MS. Strain independent variability was observed among 30 Ri-transformed root lines on the basis of lateral root density per cm (7.60 ± 0.30 to 4.5 ± 0.5), relative thickness (0.54 ± 0.07 to 1.54 ± 0.1 mm), growth index (9.16 ± 1.1 to 17.79 ± 1.35 FW basis and 10.77 ± 0.95 to 19.46 ± 1.78 DW basis) and trans-resveratrol content 0.27 ± 0.03 (root line R1000-1) to 0.969 ± 0.141 mg g DW−1 (root line RIX-19) in solid N/5 medium, which was 4.1–14 fold greater than in excised non-transformed root cultures (0.06 ± 0.01 mg g DW−1). Optimum growth and productivity in liquid culture was achieved in N/5 medium supplemented with 0.01 % activated charcoal. Root line RIX-19 showed maximum trans-resveratrol accumulation (1.21 ± 0.09 mg g DW−1) and productivity (0.37 ± 0.08 mg per flask), which was 19 fold higher than non-transformed root cultures. This optimized protocol can be utilized for large scale cultivation of transformed root cultures in industrial bioreactors for mass synthesis of trans-resveratrol. © 2015, Springer Science+Business Media Dordrecht.
Flow cytometry and its utility
(Springer India, 2017) Mihir Halder; Sayantani Nath; Sumita Jha
Flow cytometry is a laser-based biophysical technique to study optical properties of microscopic particles in fluid suspension. The technology is not only routinely used in medicine (transplantation, hematology, cancer, prenatal diagnosis, genetics, and sperm sorting for sex preselection) but also has many other applications in basic research, crop improvement, clinical practice, and clinical trials. The principles of flow cytometric chromosome analysis and sorting known as flow cytogenetics and research in this field have increased over the past four decades due to high sensitivity and precision of this technique. The use of nucleic acid-specific fluorochromes has semiautomated quantitative chromosome analysis, thus reducing subjectivity of preexisting slide-based methods. Flow cytometric classification of chromosomes (flow karyotyping) enables detection of chromosomal aberrations, while flow sorting permits isolation of single chromosome types in large quantities to be used for gene mapping, preparation of chromosome-specific gene libraries, and ultimately sequencing leading to chromosome genomics. Flow fluorescent in situ hybridization (FISH), i.e., combined fluorescent in situ hybridization in suspension (FISHIS) and flow cytometry using microsatellite DNA or gene-specific probes, is another approach to increase our knowledge on structure, function, and evolution of chromosomes. These techniques provide increased value for diagnosis and management of clinical diseases, especially cancer which is generally characterized by high chromosomal instability. Flow cytogenetics also has important applications in plant biology, especially in the study of genome evolution and crop improvement. This review outlines the utility of flow cytometry in chromosomal analysis, its applications, limitations, and future directions. © Springer India 2017. All rights are reserved.
Medicinal bulbous plants biology, phytochemistry and biotechnology
(CRC Press, 2016) Sayantani Nath; Partha Sarathi Saha; Sumita Jha
Bulbous plants are geophytes with an enlarged storage structure called bulb which helps them to perennate extreme conditions. True bulbs mostly constitute petaloid monocotyledons belonging to Amaryllidaceae, Alliaceae, Hyacinthaceae, Liliaceae and Iridaceae. These bulbous plants produce an array of unique biologically active compounds and have been extensively used in ethno medicine. Norbelladine alkaloids produced by Amaryllidaceae plants have anticancerous, acetylcholine esterase inhibitory and antimalarial activities while organosulphur compounds characteristic of the family Alliaceae are known for their antioxidant, anticancerous, antidiabetic and antihypertensive activities. Cardenolides and bufadienolides produced by members of Hyacinthaceae exhibit cardiotonic properties and have been used to treat heart failure since time immemorial. Such extensive medicinal properties of bulbous plants have resulted in their ruthless exploitation making their conservation an immediate necessity. Micropropagation, genetic improvements as well as elicitation of secondary metabolite can aid in the judicious exploitation of bulbous medicinal plants. The present review encompasses within its scope biology, phytochemistry and biotechnology of true bulbous geophytes of medicinal importance. © 2014 by Taylor & Francis Group, LLC.
Molecular phylogenetic studies based on rDNA ITS, cpDNA trnL intron sequence and cladode characteristics in nine Protasparagus taxa
(Springer-Verlag Wien, 2015) Partha Sarathi Saha; Sudipta Ray; Mainak Sengupta; Sumita Jha
The genus Asparagus comprises three subgenera of cladode bearing plants: Protasparagus, Asparagus, and Myrsiphyllum. The interspecific delimitation of the subgenus Protasparagus is ill-defined till date. In the present study, interspecific phylogenetic relationships among nine taxa of Protasparagus based on ribosomal DNA internal transcribed spacer region (ITS1-5.8S-ITS2) sequence and the chloroplast DNA trnL intron sequence conservation with their cladode morphology, anatomy, and stomatal characteristics have been analyzed for the first time. The monophyletic subgenus Protasparagus could be resolved into four strongly supported distinct subclades (I, II, III and IV) suggesting that the rDNA and cpDNA molecular phylogenies are explicitly congruent with the cladode characteristics of the subgenus Protasparagus. The present study also confirms the existing subgeneric classification of the genus Asparagus with the monophyletic origin of the dioecious subgenus Asparagus. The present work brings out phylogenetic and taxonomic relationships within the studied taxa of the subgenus Protasparagus therefore providing important background information for further studies on biogeography of a wide range of species. © Springer-Verlag Wien 2014.
Morpho-histological characterization and direct shoot organogenesis in two types of explants from Bacopa monnieri on unsupplemented basal medium
(Springer Netherlands, 2017) Sayantika Sarkar; Sumita Jha
In the present study, high frequency regeneration has been obtained via de novo direct shoot organogenesis from leaf and internode explants in Murashige and Skoog (MS) basal medium without any phytohormone supplementation in Bacopa monnieri, an indigenous traditionally used medicinal herb. Leaves and internodes from different positions were excised from 4-weeks-old in vitro propagated B. monnieri plants and cultured on MS basal medium supplemented with 3% (w/v) sucrose and 0.75% (w/v) agar for 4 weeks. The induction of de novo shoot buds was observed at petiolar cut edges of leaf and both proximal and distal cut ends of internode explants within 10–15 days of culture. The first histological changes could be observed after 4–5 days, with meristematic activity of vascular bundles. Proliferation of epidermal cells gave rise to dome-shaped protuberances followed by shoot apical meristems formation and their vascular connections with explant tissues within 2 weeks of culture. However, a basipetal gradient of shoot regeneration from both types of explants collected along the branch axis was noticed after 4 weeks of culture. Leaf and internode explants near the basal region exhibited significantly higher number of shoot buds and micro shoots (8.8/leaf explant and 15/internode explant). Microshoots (7–12 micro shoots/leaf or internode explants) elongated (shoot length 8–9 cm) within 8 weeks on phytohormone free MS medium. Excised micro shoots rooted (100%) in hormone free MS medium within two weeks of culture. Rooted plants were then acclimatized and transferred to field with 95% survival. This protocol may be used for micropropagation, genetic transformation as well as a model system for evaluation of changes associated with acquisition of competence of differentiated cells in phytohormone free medium. © 2017, Springer Science+Business Media Dordrecht.
Morphological and cytogenetical characterization of ‘Dalle Khursani’: a polyploid cultivated Capsicum of India
(Elsevier B.V., 2017) Timir Baran Jha; Partha Sarathi Saha; Sayantani Nath; Anusree Das; Sumita Jha
A morphologically different, perennial and highly pungent Capsicum cultivar ‘Dalle Khursani’ is available in the Darjeeling district of West Bengal over many decades. Its counterpart in Sikkim was identified as polyploid Capsicum cultivar with 2n = 4x = 48 chromosomes. However, cytogenetical status of the cultivar growing in West Bengal was lacking. The present study has carried out extensive morphological, chromosomal and molecular analysis on seven ‘Dalle Khursani’ populations of West Bengal. Our results on flower morphology explicitly delimit ‘Dalle Khursani’ populations from the members of ‘C. annuum complex’. Somatic and gametic chromosome analysis has clearly established their polyploid chromosome number 4x = 48 and allopolyploid nature of the cultivar (2n = 4x = 48). Fluorescent banding analysis with CMA revealed CMA + ve/DAPI-ve heterochromatin blocks in four ‘Dalle Khursani’ populations for the first time indicating the prevalence of GC rich heterochromatin in the studied taxon. RAPD based genomic DNA analysis has exhibited similar banding profile within the populations and their complete difference from the diploid members of ‘C. annuum complex’. Our present studies in conformity with our earlier report have firmly established that allopolyploidy in Capsicum is not a rarity. It is available in Indian states and will help the Capsicum researchers and breeders in their future crop improvement programmes. © 2016 Elsevier B.V.
Ribosomal DNA ITS1, 5.8S and ITS2 secondary structure, nuclear DNA content and phytochemical analyses reveal distinctive characteristics of four subclades of Protasparagus
(Wiley-Liss Inc., 2017) Partha Sarathi Saha; Mainak Sengupta; Sumita Jha
The use of ribosomal DNA (rDNA) internal transcribed spacer (ITS) primary sequence based phylogeny is a conventional practice to estimate the evolutionary interspecies relationship. However, analysis of the functional folding patterns and higher order secondary structures of ITS regions can provide additional important information regarding species relatedness and interspecies variations. In the present study, we provide the first detailed information on the rDNA ITS secondary structure diversity in the four subclades of the subgenus Protasparagus. Several angiospermic conserved motifs were identified in each of the ITS1, 5.8S and ITS2 secondary structures of the studied taxon. Topological comparison of the ITS1 secondary structures showed variations in the helix- IV regions. Moreover, presence of unique sequence motifs and differences in the internal loop structures were found to be subclade specific. The present study suggests that comprehensive analysis of the ITS1, 5.8S and ITS2 structural elements including helices, loops and bulges can be used as an important tool for species delimitation. The present study investigated the evolution of the secondary structure of ITS marker (its phylogenetic utility), genome size, base chromosome number and phytochemicals, and identified a putative polyploid event shared by a number of Protasparagus species. The phytochemical analysis of two important active compounds, i.e., shatavarin-IV and sarsasapogenin, also reveals their presence in all the studied taxa constitutively even at the subclade level. © 2016 Institute of Botany, Chinese Academy of Sciences