Cloning And Function Analysis Of ESP2, A Gene Involving Enclosed Short Panicle In Rice

Genetically eliminating the panicle enclosure phenomenon in male sterile lines has been an important goal in hybrid rice breeding. For this purpose, it is necessary to research in depth the genetic cause of panicle enclosure. We found a new enclosed panicle mutant, named as esp2 (enclosed shorter panicle 2), from the tissue culture progeny of indica rice cultivar Minghui-86. Previous studies indicated that the mutant resulted from a recessive mutation in a single gene, and fine mapped the gene into a 14-kb region on chromosome 1. Based on these results, this study aimed to identify the candidate gene of ESP2 and validate the function of the candidate gene by genetic complementation test. The main results are as follows:1. Gene prediction indicated that only one intact gene exists in the fine mapped 14-kb region, namely, a putative phosphatidylserine synthase (putative phosphatidylserine synthase, PSS) gene. Sequencing analysis suggested that the 4th exon of the PSS gene was inserted by a 5287-bp retrotransposon. Therefore, we took this gene as the candidate of ESP2.2. Analysis of promoter region of the ESP2 candidate gene using the software BDGP indicated that conserved promoter elements exist within the 2 kb region and the 2 kb-4 kb region upstream of the start codon. To identify the function of the ESP2 candidate gene, two vectors for complementation test, pCAMBIA1301-ESP2-S and pCAMBIA1301-ESP2-L, were constructed, which carried a 1952-kb and 3537-kb promoter sequence of the ESP2 candidate gene, respectively. The two vectors were transfered into the calli induced from immature embryos of the mutant, and 56 and 16 transgenic plants were obtained, respectively. The transgenic plants looked similar to the wild type at the heading stage without the phenotype of enclosed panicle. Therefore, the PSS gene should be ESP2.