MOLECULAR CHARACTERIZATION OF NBS-LRR RESISTANT GENE ANALOGUES (RGAs) FROM INDIGENOUS AND WILD BANANA (MUSA SPP.) GERMPLASM
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Date
2018-01
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AAU, Jorhat
Abstract
Commercial banana varieties are highly susceptible to fungal and bacterial
pathogens, nematodes, viruses and insect pests. The largest known family of plant
disease resistance (R) genes encodes proteins with nucleotide-binding site (NBS) and
leucine-rich repeat (LRR) domains. Conserved motifs in such genes in diverse plant
species offer a means for the isolation of candidate genes in banana that may be
involved in plant defense. In the present study, an attempt was made to isolate and
characterize the conserved region NBS of the NBS-LRR resistance gene analogues
(RGAs) from locally cultivated indigenous and wild banana germplasms of Assam.
The investigation was started with the isolation of genomic DNA from ten cultivated
indigenous germplasms viz. Kach kol, Cheni champa, Ukho jahaji, Malbhog,
Manuhor, Athiya kol, Bhim kol, Ketekihunda, Phesa manuhor and Ximolu manuhor
and, five wild germplasms (designated as W1, W2, W3, W4 and W5). To target the
NBS region of the banana germplasms, four pairs of PCR primers out of which two
were degenerate primers, were designed from existing NBS-LRR sequences available
in the GenBank. After successful isolation and sequencing of the PCR amplified NBS
fragments from all the fifteen samples, confirmation about the identity of the
sequences was done by homology search using BLASTn and BLASTp algorithms
which revealed the sequences to be significantly similar to the NBS-LRR class
disease resistance proteins available in NCBI. The sequence identity was further
confirmed by checking for the Pfam NB-ARC domain, which is a protein domain
characteristic of the plant resistance NBS-LRR protein. The NB-ARC domain was
obtained in all the isolated NBS sequences. Finally, the presence of the consensus
sequence for Kinase-2 motif (LLDDVW) and phylogenetic analysis of the isolated
NBS sequences further provided evidence that the sequences belong to the typical
non-Toll/interleukin-1 receptor- like domain NBS-LRR gene family, as expected. As
a future prospect, upon cloning of the full length NBS-LRR sequences from these
germplasms would open up possibilities for development of disease resistant cultivars
through genetic engineering approaches.