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为了克隆DL-6基因,以dl-6与9311配制正反杂交组合进行性状分析发现,dl-6突变性状受1对隐性核基因控制,利用SSR分子标记将控制dl-6性状的基因DL-6初步定位在水稻第3染色体的短臂上,进一步利用新发展的In Del标记将DL-6基因精细定位在I3-5和I3-8之间85 kb的物理距离内。对该区段内存在的开放阅读框(ORF)进行分析,发现其中ORF9编码的YABBY基因是一个与叶脉发育相关的基因,对突变体dl-6和野生型中YABBY基因进行测序,将测序结果与数据中日本晴序列进行比对发现,突变体dl-6中YABBY基因的第1个外显子存在1个单碱基突变,该突变导致野生型中编码的半胱氨酸突变为突变体中的精氨酸;同时突变体dl-6在YABBY基因的3’端还存在8个碱基的缺失。这2个突变位点哪个是导致dl-6突变的功能区目前还不确定,有待进一步研究。
In order to clone DL-6 gene, dl-6 and 9311 were used to prepare the positive and negative hybrid combinations for trait analysis. The results showed that the dl-6 mutation was controlled by one pair of recessive nuclear genes. SSR markers were used to control the gene DL- -6 was initially located on the short arm of chromosome 3 of rice and the DL-6 gene was further finely mapped within a physical distance of 85 kb between I3-5 and I3-8 using the newly developed In Del marker. The open reading frame (ORF) existed in this region was analyzed and found that the YABBY gene encoded by ORF9 was a gene related to vein development. The YABBY gene in mutant dl-6 and wild type was sequenced. The sequencing results In comparison with the Nipponbare sequence data, there was a single base mutation in the first exon of the YABBY gene in mutant dl-6, which resulted in the mutation of the cysteine encoded in the wild type into the mutant Arginine; At the same time mutant dl-6 in the 3 ’end of the YABBY gene there is also the deletion of 8 bases. Which of the two mutation sites is the cause of dl-6 mutation in the functional area is still uncertain, pending further study.