山羊草属杀配子染色体2C特异SCAR标记的建立

用40条10碱基随机引物,对普通小麦“中国春”、“中国春”-杀配子染色体2C二体附加系、柱穗山羊草进行RAPD分析,筛选到1个杀配子染色体2C特异引物OPF03。从“中国春”-杀配子染色体2C二体附加系中克隆了该DNA片段。测序结果表明,该片段长1 496 bp,与大麦1个cDNA的同源性为92%。根据OPF03₁₄₉₆的序列设计了1对SCAR引物2C-F₅₈₆、2C-R₅₈₆,对普通小麦“中国春”、“中国春”-杀配子染色体2C二体附加系、柱穗山羊草、二倍体长穗偃麦草、“中国春”-长穗偃麦草7E二体附加系等材料进行了SCAR分析,在“中国春”-杀配子染色体2C二体附加系、柱穗山羊草中扩增出了586 bp的片段,而在普通小麦“中国春”、二倍体长穗偃麦草、“中国春”-长穗偃麦草7E二体附加系中没有该产物,初步证明此标记可以用于杀配子染色体2C的检测。进一步用该对SCAR引物对“中国春”-杀配子染色体2C二体附加系与“中国春”-长穗偃麦草7E二体附加系的杂种F₁代、F₂代进行了分析,结果在全部F₁代以及部分F₂代材料扩增出了目的片段,进一步证明了此SCAR标记可以用来检测小麦背景下的杀配子染色体2C,为小麦背景中杀配子染色体2C的快速跟踪检测提供了新途径。

Establishment of a gametocidal chromosome 2C specific SCAR marker

A total of 40 decamer oligonucleotides were used as primers to perform random amplified polymorphic DNA (RAPD) analysis on common wheat CS (Chinese spring), CS-2C disomic addition line, Aegilops cylindrica. Of these primers, one could amplify a specific DNA fragment in CS-2C disomic addition line and A. cylindrica. The fragment OPF03₁₄₉₆ was cloned and sequenced; Sequence data searching in GenBank showed that this fragment was 92% homologous to a cloned cDNA sequence from Barley. Based on the sequence of OPF03₁₄₉₆, a pair of SCAR (sequence characterized amplified region) primer 2C-F₅₈₆, 2C-R₅₈₆ was designed. SCAR analyses were performed on CS, CS-2C disomic addition line, A. cylindrica, and Thinopyrum elongatum (2X) CS-7E disomic addition line. The amplification results of SCAR primer showed that the SCAR₅₈₆ marker appeared in the CS-2C disomic addition line and A. cylindrica but not in CS, T. elongatum (2X) CS-7E disomic addition line. Furthermore, SCAR analyses were also performed on F₁ and F₂ progenies between CS-2C and CS-7E disomic addition lines. PCR amplification results showed that the SCAR₅₈₆ marker was in all of the F₁ progenies between CS-2C and CS-7E and in some of the F₂ progenies. The SCAR₅₈₆ marker can be used for detecting the gametocidal chromosome 2C in common wheat backgrounds.