Cloning and Molecular Evolution Analysis of NBS Class Resistance Gene Analogs in Black Bamboo (Phyllostachys nigra)
Abstract The nucleotide binding site (NBS) domain sequences were isolated from genomic DNA in black bamboo, using the degenerate primer designed according to the conserved motifs of the NBS resistance gene. The expected size of the PCR product was about 700 bp. Among 55 positive clones, the amino acid sequence alignment identified 33 black bamboo resistance gene analogs (RGAs) that contain the NBS conserved motifs. All of the 33 RGAs ORFs were constructed in an NJ (Neighbor-joining) tree, and divided into 10 groups. This analysis demonstrated the diversity of the NBS class RGA in black bamboo. The maximum likelihood estimates of various evolutionary models were analyzed; the result showed that 2 groups with a total of 10 sequences and 12 sites demonstrated statistically significant positive selection. Most of the positive selected sites were not located in the NBS conserved motifs. Two groups of gene conversion events had been discovered, which provide a material basis and research direction in isolating black bamboo R genes.
