摘要
采用Dynal磁珠富集法构建了香椿微卫星富集文库,通过测序结果对文库的特性进行了分析。实验使用改良CTAB法提取香椿基因组DNA,用（AC）8、（AG）8和（ATG）123种带有生物素标记的探针与香椿基因组DNA的酶切片段进行杂交,将磁珠捕获的含有微卫星序列的DNA片段插入p MD 18-T载体,并转入感受态细胞Trans 5α构建克隆,经筛选后得到含356个克隆的香椿基因组微卫星富集文库。从富集文库中挑选插入片段长度为400-800 bp的128个克隆进行测序,其中含有SSR的序列77条,得率达60.16%,其中完美型占82.69%,非完美型8.65%,混合型8.65%。上述结果为SSR位点的进一步开发奠定了基础。
Toona sinensis is peculiar economic tree species in China,people like eating itg early spring bud as vegetables. This research used the method of Dynal magnetic beads enrichment to build enriched microsatellite sequence library, analysed the characteristics of library through sequencing results. Total genomie DNA of Toona sinensis was extracted using the modified CTAB method, the digested Toona sinensis genomic DNA fragments were hybridized with the biotinylated probes （ AC ） s, （AG） s and （ATG） 12. The enriched DNA fragments were ligated into pMD 18-T easy vector and then transformed into competent cell Trans 5 a to form an enriched microsatellite sequence library which has 356 clones. Reverse as primers identified 128 clones containing 400- 800 bp inserts. Sequence analysis of these positive clones confirmed 77 microsatellite sequences, with an enrichment efficiency of 60.16%. Of these microsatellite loci ,82.69% were perfect repeat motif,8.65% imperfect and 8.65% compound repeat motifs. The results lay the foundations for the further development of SSR loci.
出处
《种子》
北大核心
2015年第2期13-16,23,共5页
Seed
基金
国家自然科学基金项目(编号:31170351)
河南省教育厅自然科学基金项目(编号:2008 A 180012)
关键词
香椿
微卫星
磁珠
富集文库
Toona sinensis
microsatellite
magnetic beads
enriched library
