基于寡核苷酸探针套painting的中国春非整倍体高清核型及应用

基于寡核苷酸探针套painting的染色体鉴定技术简单、经济和高效, 可以促进小麦品种及亲缘物种染色体识别和变异体鉴定, 提高染色体工程效率。我们前期开发了寡核苷酸探针套, 包含pAs1-1、pAs1-3、AFA-4、(GAA)₁₀和pSc119.2-1共5个探针。本研究通过一次荧光原位杂交(FISH), 对源于17个非整倍体的18份材料分析发现, 其中14个染色体组成正确, 可以清晰识别相应的缺体、四体和端体。还构建了基于寡核苷酸探针套涂染的、能准确识别3个基因组和7个部分同源群染色体的高清核型, 发现4个非整倍体发生变异, 其中从N5BT5D中鉴定出一个可能的小片段相互易位系T6AS·6AL-6DL和T6DS·6DL-6AL。进一步对7个地方品种、10个栽培品种(系)和1个人工合成小麦分析, 发现15条染色体存在多态性, 涉及6条B组(除4B)、5条A组(除1A和3A)和4条D组(1D、2D、4D和7D)染色体, 可以清晰识别我国小麦生产上广泛应用的3种易位类型(T1RS·1BL、T6VS·6AL及相互易位T1RS·7DL和T7DS·1BL), 省去了基因组原位杂交(GISH)程序。另外, 对5个亲缘物种分析发现, 该探针套可以识别栽培一粒小麦、硬粒小麦Langdon、荆州黑麦、长穗偃麦草(2n=2x=14)全部和中间偃麦草30条染色体, 并构建了这5个物种的核型。本研究结果证实该寡核苷酸探针套可以有效用于小麦及亲缘物种染色体鉴定, 高清晰的中国春非整倍体核型为小麦染色体工程提供了参考标准。

Development and Application of High Resolution Karyotypes of Chinese Spring Aneuploids

Oligonucleotide (oligo hereafter) multiplex-based chromosome painting facilitates chromosome identification of both wheat cultivars and its relatives in a simple, easy and high efficient way. In this study, an oligo multiplex containing oligos pAs1-1, pAs1-3, AFA-4, (GAA)₁₀, and pSc119.2-1 developed earlierwas used for chromosome painting of 18 accessions from 17 Chinese Spring (CS) aneuploids. The high resolution karyotypes allowed to clearly distinguish individualwheat chromosomes. Fourteen aneuploidshad theexpected chromosome constitutions whereasthe other four had chromosome variations including one witha possible small segmental reciprocal translocation T6AS·6AL-6DL and T6DS·6DL-6AL occurred in N5BT5D. The following analysis on eightlandraces,ninecultivars (lines), and one synthetic hexaploid wheat, observed karyotypediversities from 15 chromosomes including six B- (except for 4B), five A- (except for 1A and 3A), and four D-genome (1D, 2D, 4D, and 7D) chromosomes. The three widely-used translocations in China, i.e. T1BL·1RS, T6AL·6VS and the reciprocal translocation T1RS·7DL and T7DS·1BL, were clearly detected after only oncefluorescence in situ hybridization(FISH) using the oligo multiplex and without genomic in situ hybridization (GISH). This oligo multiplex also produced rich signals in all chromosomes of Triticum monococum, rye cultivar ‘Jingzhouheimai’, durum wheat ‘Langdon’, and Thinopyrum elongatum, and 30 chromosomes of Thinopyrum intermedium. The karyotypes of these five species were thus developed. These results indicate that oligo multiplex-based chromosome painting will play active roles on chromosome identifying, and provide a reference for the standard karyotypes of CS aneuploids.