NISCAIR Online Periodicals Repository Collection: IJBT Vol.09(2) [April 2010]

Development, micropropagation and characterization of colchiploid of Echinacea purpurea (L.) Moench

Mon, 29 Mar 2010 22:58:59 GMT

Title: Development, micropropagation and characterization of colchiploid of Echinacea purpurea (L.) Moench

Authors: Koul, Sushma; Sambyal, Manju; Kitchlu, S K; Bakshi, S K; Kaul, M K

Abstract: To enhance the genetic resource base of Echinacea purpurea (L.) Moench, a tetraploid was developed by treating seeds with different concentrations of colchicine. For further multiplication, in vitro micropropagation through adventitious shoot induction and proliferation was achieved. Shoot buds were induced directly on adaxial surface of mature leaf tissues growing on MS medium supplemented with equimolar concentration of BAP and IBA. The optimum initial shoot regeneration frequency (63%) with average 2.3 shoots per explant was achieved after 30 d of culture initiation. Maximum shoot organogenesis, with 8-10 shoots per culture flask, was obtained on the same medium after 10^th subculture. Proliferating callus with intermixed shoot buds was derived from leaf tissue explants placed on MS medium supplemented with 1 mg L^-1 each of BAP and IAA. The regenerated shoots were rooted on MS medium devoid of any growth regulator. Rooted plantlets were hardened under greenhouse conditions at 20-22°C with 85-90% RH. About 95% plantlets survived acclimatization, producing phenotypically normal plants in the greenhouse. After 4 wks, rooted plants were successfully transferred to soil in experimental field station at Jammu. Comparable field trials were laid with parental diploids. Tetraploids were dwarf, showed low seed set but were superior because of higher caffeic acid content. The results of this study have established a procedure for development and propagation of tetraploid of E. purpurea having higher caffeic acid content.

Page(s): 221-224

An efficient protocol for in vitro regeneration of Podophyllum hexandrum, a critically endangered medicinal plant

Mon, 29 Mar 2010 22:58:59 GMT

Title: An efficient protocol for in vitro regeneration of Podophyllum hexandrum, a critically endangered medicinal plant

Authors: Chakraborty, Amrita; Bhattacharya, Dipto; Ghanta, Srijani; Chattopadhyay, Sharmila

Abstract: Rhizome explants were used to develop a protocol for in vitro plantlet regeneration of Podophyllum hexandrum, a critically endangered medicinal plant, through direct organogenesis. Highest rate of multiple shoot formation was noted in MS medium supplemented with 11.42 μM IAA within three months. A synergistic effect of 2.68 μM NAA and 11.1 μM BAP was second best. The ½ MS liquid medium with 100 μM IBA was most suitable for rooting of shoots. Leaf explants resulted in callus formation only. Leaf regenerated calli showed the presence of podophyllotoxin in HPLC analysis.

Page(s): 217-220

An efficient genomic DNA isolation protocol for RAPD and SSR analysis in Acorus calamus L.

Mon, 29 Mar 2010 22:58:59 GMT

Title: An efficient genomic DNA isolation protocol for RAPD and SSR analysis in Acorus calamus L.

Authors: Ginwal, H S; Mittal, Neha

Abstract: Molecular based genetic analysis requires good quality of DNA in sufficient quantity to generate robust DNA fingerprints. The leaves of sweet flag (Acorus calamus L.) contain high amounts of polyphenolic compounds and polysaccharides, which interfere in the amplification reactions. Four genomic DNA extraction methods were tested for yield, quality and suitability of genomic DNA for RAPD (random amplified polymorphic DNA) and SSR (simple sequence repeat) marker analysis in sweet flag. Fresh young leaves were subjected to the already available protocols and a procedure was devised after modification in the protocol of Stange et al for the isolation of total genomic DNA. The modified method employs high concentrations of polyvinyl pyrrolidone, addition of lithium chloride solution as well as additional washing step of DNA pellets. The yield was found approximately 200-500 µg DNA per 100 mg of plant material and the purity ratio was found 1.7-2.0. Usage of highly concentrated PVP extraction buffer and chloroform:isoamylalcohol repetition step was found useful to overcome problems from polyphenolic compounds and polysaccharides. The extracted DNA through this method was found suitable for RAPD and SSR analysis.

Page(s): 213-216

Use of in silico and semiquantitative RT-PCR approaches to develop nutrient rich rice (Oryza sativa L.)

Mon, 29 Mar 2010 22:58:59 GMT

Title: Use of in silico and semiquantitative RT-PCR approaches to develop nutrient rich rice (Oryza sativa L.)

Authors: Banerjee, Shubha; Sharma, D J; Verulkar, S B; Chandel, G

Abstract: Inspite of striking agricultural progress and adequate food grain production, protein energy and micronutrient malnutrition are widespread among rural and poor population. The pharmaceutical and diet diversification based approaches have achieved little success due to poverty and educational unawareness. Biofortification of staple food crop rice, which is consumed in large amounts daily, will serve as an important vector to combat malnutrition. The possible approach to improve nutritive value of rice involves exploitation of available genetic variability for grain protein and Fe/Zn contents with conventional and modern biotechnological tools. We have analysed variability in grain protein and Fe/Zn levels in rice, and factors affecting them for identification of rice genotypes with higher nutritive value. Wide variation for grain protein and micronutrient levels were recorded among the tested rice genotypes, which ranged from 6.19 to 10.75% for grain protein content, 4.82 to 22.69 μg/g for grain Fe and 13.95 to 41.73 μg/g for grain Zn content. Significant effect of nitrogenous fertilizer dose and native soil properties were observed on grain protein content, whereas grain Fe and Zn levels were more significantly affected by native soil properties and showed non-significant effect of nitrogen applied. Three genotypes, R-RF-31, Lalmati and R 1033-968-2-1 were identified as high protein and Fe/Zn containing rice lines. In view of our findings and previous studies, where significant Gene Environment (G E) effect has been reported on grain nutritive traits, molecular markers are of great use for intensive screening of large populations and identification of environmentally stable outperforming genotypes. The genomic sequence of candidate genes governing micronutrient content in rice were, thus, analysed in silico for identification of novel gene specific markers (SSRs and SNPs) and expression sequence tags (ESTs and MPSS) to understand putative expression pattern. Further, to confirm in silico expression results and functionally characterized Fe and Zn homeostasis related genes, root and shoot transcriptome analysis of a set of 12 diverse rice genotypes was carried out for expression of 21 metal homeostasis related genes belonging to OsYSLs, OsFROs, OsZIPs, OsNRAMPs and OsFERs families, and OsNAAT1, OsVIT1, OsNAC and OsNAS2 genes. A total of 176 novel SNPs and 39 novel SSRs were identified within metal related genes, which can be used for developing gene specific markers. The ESTs and MPSS tag based in silico expression analysis results were in consistency to the semi-quantitative RT-PCR based transcriptome analysis. Expression profiling of rice root and shoot transcriptome at maximum tillering and mid-grain filling stages revealed high level of expression of most of the Fe and Zn uptake and transport related genes in genotypes showing higher grain Fe and Zn concentrations. The rice genotypes with high grain protein and micronutrients, identified in this study, will provide the basis of bioavailability assay and will also serve as potential genetic material for molecular breeding of nutrient rich rice.

Page(s): 203-212