Development of molecular markers in weedy rice chloroplast based on genomic database.

[Objectives]The origin and evolution of weedy rice still is a disputed but hot issue.The molecular markers applied in a study of weedy rice are limited and not specific to weedy rice discriminating from cultivated rice.Hence,it is necessary to explore new molecular markers in this field.[Methods]The downloaded chloroplast genome sequences of 13 cultivated rice and wild rice from genomic database were aligned and analyzed.We designed primers as markers based on the sequence of the high frequency variation regions.Then the labeling areas in 60 accessions of weedy rice were sequenced and verified for the polymorphism.[Results]The results showed that there were 1 020 SNPs and 111 In Dels in the chloroplastic genome sequences of 13 cultivated rice and wild rice,which had rich genetic variation.Three primers were designed as markers in the selected 3 high frequency variation regions which were located in trn G-trnf M inter spacer,psb M-trn C inter spacer and trn T-trn L inter spacer.The detection of 60 weedy rice accessions using the developed three markers demonstrated that there were 7 SNPs and 6 In Dels in these three regions with a total sequence of 1 966 bp among the weedy rice.The total length of the variation sequence contained 34 bp with 1.73% variation of the whole sequence length.The phylogenetic tree constructed by tag sequence divided the 60 weedy rice accessions into four groups.Additionally,taking 13 cultivated rice and wild rice sequences from the network database,it was found that japonica and indica rice and cultivated rice were always clustered into the corresponding weedy rice classes.[Conclusions]The three markers developed in this study could well classify weedy rice accessions and showed a potential application for weedy rice genetic diversity and evolution research.