菠菜性别差异NUPTs序列的克隆及分析

质体DNA向核基因组转移能够形成核质体DNA,核质体DNA序列的插入是推动动植物基因组及染色体组演化的重要动力.但是核质体DNA的插入和植物性染色体起源及演化之间的关系仍不清楚.以雌雄异株植物菠菜为材料,利用基因组消减杂交技术筛选分离菠菜雌雄基因组差异的NUPTs序列,并进行验证和分析.结果表明,从构建的菠菜消减杂交文库中共得到39条长度为75~308 bp的雌雄差异序列,其平均长度约为154 bp.对获得的序列进行Blastn同源比对发现了12条序列为叶绿体基因组来源序列,这些序列与菠菜叶绿体基因组相似度在98%以上,说明所获得的差异片段为核质体DNA序列.将筛选出的核质体DNA序列进行进一步验证后获得了两个稳定的长度分别为146 bp和199 bp的雄性偏好核质体DNA序列,说明所获得的两个NUPTs序列在菠菜雄性基因组中有更多的累积.

Cloning and Analysis of Sex-biased NUPTs in Spinach

The transfer of plastid DNA into nuclear genome leads to the formation of nuclear plastid DNA( NUPT). The insertion of NUPTs is an important force to drive the evolution of genome and chromosomes both in animals and in plants. However,it is still unclear about the relationship between the insertion of NUPTs and plant sex chromosome origin and evolution. In this study,sex different NUPT sequences were screened and isolated using genome substractive hybridization technique,and further validation and analysis were also conducted. A total of 39 valid sequences with the length between 75 bp and 308 bp( average of 154 bp) were obtained from the spinach substractive hybridization library. Blastn searching revealed that 12 sequences were related to chloroplast DNA,showing over 98% similarity with spinach chloroplast DNA. The results indicated that these 12 sequences were NUPTs in spinach genome. Finally,two stable male-biased NUPTs with size of 146 bp and 199 bp were obtained by further genomic semi-quantitative PCR validation,demonstrating that these two NUPTs accumulated more in spinach male genome than in female genome.