中间偃麦草第6同源群特异STS标记开发

中间偃麦草(2n=6x=42,JJJ^sJ^sStSt)是小麦遗传改良的重要野生资源之一,因其基因组尚未完成测序,造成目前已报道的特异分子标记数量较少,不能满足小麦生产和研究领域内对杂交材料中外源片段或外源基因鉴定的需求。本研究利用中间偃麦草GBS芯片探针数据组装了5877409条Contig序列,筛选后获得5452条与小麦基因组相似性低于80%的、具有染色体位置信息的非冗余序列,据此开发2019个序列标签位点(sequence-tagged site,STS)分子标记,在中间偃麦草第1至第7同源群中的分布依次为250、215、323、253、323、253和402;利用5株中间偃麦草和5份小麦农家种的DNA从253个第6同源群(G6)标记中进一步筛选出160个中间偃麦草特异标记,其中"+/-"型特异标记共有53个,分布在G6-Chr1(32个)、G6-Chr2(13个)和G6-Chr3(8个)染色体上;接着利用拟鹅观草(2n=2x=14,StSt)、百萨偃麦草(2n=2x=14,J^bJ^b)、二倍体长穗偃麦草(2n=2x=14,J^eJ^e)以及中国春-二倍体长穗偃麦草6J^e代换系推断G6-Chr2为6St染色体;最后利用6St上开发的13个"+/-"型标记,从分子水平对小麦-偃麦草代换系F881(6St/6D)中的外源6St染色体进行了验证。研究结果将为中间偃麦草染色体或染色体片段的鉴定提供较为方便和经济的检测手段。

Development of specific STS markers for the sixth homologous group of Thinopyrum intermedium

Thinopyrum intermedium (2n=6x=42, JJJ^sJ^sStSt) is one of the important wild resources for wheat genetic improvement. However, as the Th. intermedium genome has not yet been sequenced, the numbers of reported Thinopyrum-specific molecular markers are insufficient for the purpose of identification of the small aline fragments or genes in the offspring of wheat-Th. intermedium hybrids in wheat breeding research. In this study, a total of 5877409 Contigs (overlapping DNA sequences) were obtained by assembling the probe sequences of a Th. intermedium genotyping-by-sequencing chip downloaded from a database. Through informatics analysis, 5452 non-redundant Contigs with chromosome location information and a similarity less than 80% with the wheat genome were screened out, and 2019 sequence-tagged site (STS) molecular markers were developed. The distribution numbers of markers from Thinopyrum homologous groups 1 (G1) to G7 were 250, 215, 323, 253, 323, 253 and 402, respectively. Then, 160 Thinopyrum-G6 specific markers were selected based on amplification of five Th. intermedium plants and five wheat landraces, 53 of which were of “+/-” specificity and were distributed on the three G6 chromosomes: G6-Chr1 (32), G6-Chr2 (13) and G6-Chr3 (8). Next, G6-Chr2 was deduced to be the 6St chromosome by using three progenitor species of Th. intermedium, including Th. elongatum (2n=2x=14, J^eJ^e), Th. bessarabicum (2n=2x=14, J^bJ^b) and Pseudoroegneiria strigosa (2n=2x=14, StSt), as well as 6J^e substitution lines of Chinese spring-Th. elongatum. Finally, the 13 “+/-” specific markers developed that were located on Thinopyrum-6St were used to verify the exogenous chromosome in wheat-Thinopyrum substitution line F881-6St/6D at the molecular level. These results will provide a more convenient and economical method for the identification of chromosomes or chromosome fragments of Th. intermedium.