Prashant Shekhar Gupta

Inter simple sequence repeat profile as a genetic marker system in sugarcane

Molecular profiling for varietal differentiation assumes an important role in the light of UPOV initiatives and PVP act. Inter simple sequence repeats (ISSR), a PCR based genetic marker system circumvents the requirement for flanking sequence information, and thus has found wide applicability in a variety of plants. The potential of ISSR markers for molecular profiling was assessed in sugarcane using a panel of 42 varieties of subtropical India. ISSR amplification was achieved using di-, tri-and tetraoligonucleotide repeats. All the nucleotide motifs were amenable to amplification. The ISSR results suggested that the frequency and clustering of specific simple sequence repeats was variable and motif-specific, however, there was not much difference in the di-, tri-or tetra-nucleotide ISSR profiles. The ISSR amplification pattern was used to assess genetic similarity of these sugarcane genotypes by Dice similarity coefficients, which ranged from 0.68 to 0.97 with a mean similarity value of 0.878. The ISSR amplification proved to be a valuable method for determining genetic variability among sugarcane varieties and for identification of the cultivars.

Assessment of genetic purity of micropropagated plants of sugarcane by isozyme and RAPD analysis

Shoot tip expiants of nine sugarcane varieties(Saccharum species hybrid.), CoLk 8102, CoLk 8001, CoS 767, CoLk 9606, CoLk 9617, CoS 95255, BO 91, CoJ 64 and Co 1148, were established on a solid MS medium containing 0.8% agar, 3% sucrose and 0.05mg/l of IAA, BAP and kinetin. Shoot regeneration has been achieved in liquid MS medium containing 0.5mg/l each of IAA, BAP and kinetin. Multiple shoots were obtained on liquid MS medium containing 0.01mg/l IAA, 0.5mg/l each of 6- benzyl aminopurine (BAP), kinetin and rooting occurred in 1/2 strength liquid MS medium with 5.0 mg/l NAA and 0.01 mg/l each of BAP and kinetin. To detect the genetic purity of in vitro raised plantlets, expression of iso-peroxidases in donor plants andin vitro raised plantlets was compared through native PAGE. Different genotypes showed marked variation regarding number, position and intensity of bands. Micropropagated plantlets showed identical pattern of peroxidases to their parental clones. The genetic stability of micro propagated plantlets was also tested by RAPD analysis. The banding pattern of PCR amplified products from micro propagated plantlets was monomorphic based on RAPD profile in all the varieties. RAPD analysis and peroxidase isozyme pattern confirmed the genetic purity of sugarcane plantlets derivedin vitro.

SDS and Native page protein profile for identification and characterization of elite sugarcane genotypes

The genetic markers provided by protein and enzyme polymorphism in plant population have been used to assess the degree of genetic variability and species relationship. Electrophoretic protein analysis holds potential utility in breeding programmes of sugarcane through helping in the unequivocal characterization of varieties used. Nine sugarcane cultivars were characterized by analyzing protein polymorphism in leaf tissues through SDS and Native PAGE. The electrophoregram showed a total of twenty bands of SDS proteins with molecular weight from 10 to 200 KDa and twenty four bands of native proteins having molecular weight of 10 to 708 KDa with difference in number of bands, bandwidth and intensity among different genotypes of sugarcane. The cultivars could be differentiated on the basis of SDS and Native protein polymorphism of total soluble leaf proteins profile which showed genetic homology as well as divergence among the different genotypes. The protein gel electrophoresis approach could thus be defined as a valuable means for distinguishing accessions of sugarcane through detecting their polymorphism.

Ribosomal DNA Sequence Based Characterization of Trichoderma Isolates Antagonistic to Colletotrichum falcatum Causing Red Rot Disease of Sugarcane

Trichoderma bioagent isolates cultured from the sugarcane rhizosphere of subtropical India, and established efficient against Colletotrichum falcatum Went (red rot pathogen) in sugarcane are described based on phylogenetic analysis of nucleotide sequences. DNA sequences of the isolates included the internal transcribed spacer (ITS) regions of the rDNA cluster (ITS1-5.8S- ITS2). The 5.8S region was found conserved and much of the sequence variability was due to indels or transition/transversion mutations in the ITS1 and 2 regions. Both the isolates belonged to the T. harzianum clade. These isolates are distinct yet have close genetic similarity with several established strains/isolates of T. harzianum reported worldwide. Correct identification of these bioagents would augment the effective utilization of these fungi of immense agricultural importance in managing the red rot disease of sugarcane.