Sequencing Of Chloroplast DNA Genomes Of Miscanthus Sinensis & M.Sacchriflorus(Poaceae) And Development Of CpInDel Markers

Miscanthus Andersson (Poaceae) is a type of tall perennial grass with a C4 photosynthetic pathway. It is considered as one of the most promising bioenergy plants in the world because of its high biomass production, extensive adaptability and strong stress tolerances. Compared to the nuclear DNA genome, chloroplast DNA genome is highly conserved during the evolution and therefore is a kind of ideal molecular marker for studying the systematics and evolution at different taxonomy levels. Sequencing Miscanthus chloroplast DNA genome and developing chloroplast DNA markers can play an important guiding role for Miscanthus taxonomy correction, cross breeding or chloroplast genetic engineering. The main results in this study were as follows:1. The chloroplast whole DNA genomes of M. sinensis (C0430) and M. sacchariflorus (A0117) were sequenced using the whole-genome shotgun sequencing strategy and Illumina Hiseq2000 sequencing system. Based on the obtained draft whole genome sequences of these two species, the gaps in genome were corrected using PCR method and DNA fragment sequencing. The chloroplast DNA genome of M. sinensis and M. sacchariflorus was with a length of 141,369 bp and 141,133 bp, respectively, and from which about 337 chloroplast SSR loci and 50 chloroplast InDel loci were identified.2. The primer pairs for 12 chloroplast InDel loci were developed following their flanking sequences. They can effectively amplify from 43 Miscanthus genotypes and 2 Sorghum germplasms as the outgroup. Of those, five DNA markers were species-specific. The further UPGMA clustering analysis indicated that there was a closer genetic relationship between M. sacchariflorus and M. lutarioriparius, as well as between M. sinensis and M. floridulus. The InDel markers that were developed in this study showed a practical value in studying Miscanthus taxonomy.