芦笋超雄株STS双分子标记的开发及应用

Development and Application of STS Bimolecular Markers for Asparagus Super-male Plants

芦笋超雄株的获得是芦笋全雄系育种的关键,但是传统的全雄系材料需要通过杂交后代的分离情况来鉴定,极大增加了育种的周期和成本。因此,设计和开发芦笋超雄株植株性别鉴定标记具有重要的实践价值。本研究首先构建了968株芦笋两性株自交群体。在此基础上,根据Y染色体特异区基因SOFF和X染色体特异区基因WIP2/NTT设计开发了性别鉴定的STS双分子标记YSM和XSM。通过对单株性别鉴定结果表明,该STS双分子标记检测后出现293 bp单一条带的为超雄株,共有58株;出现245 bp单一条带的为雌株,共有550株;而出现245 bp和293 bp双条带的为雄株,共有360株,分别将293 bp和245 bp大小的条带进行测序后发现,两个序列和设计引物所用目的序列一致性均为100%。对开花后鉴定为雄株的418株单株逐一测交,对测交组合F_(1)的雌、雄性鉴定分析,发现有58个测交组合F_(1)全部为雄株,证明他们的父本为超雄株;有360个测交组合F_(1)出现雌雄株分离,证明他们的父本为雄株。这和STS分子标记鉴定结果完全一致,说明本研究的STS双分子标记可以准确鉴定芦笋雌株、雄株及超雄株性别。这为育种者进行芦笋全雄分子标记辅助育种提供了强有力的技术支撑。

Identification of super-male plants is the key to all-male breeding in asparagus,while traditional method based on analyzing the separation ratio of hybrid offspring remains time-consumption and cost. It is of practical value to design and develop sex-specific markers assisted for asparagus super-male plant selection. In this study,968 selfing populations of andromonoecious plants were constructed. STS bimolecular markers YSM and XSM,which can amplify the Y chromosome specific region gene SOFF and the X chromosome specific region gene WIP2/NTT,respectively,were produced. Based on the genotyping result using STS marker,58super-male plants carrying a single 293 bp fragment,550 female plants carrying a single 245 bp fragment,as well as 360 male plants with two fragments(245 bp and 293 bp)were identified. Sanger sequencing of the 293 bp and 245 bp fragments revealed a sequence identity to the reference. By making crosses using 418 male plants,58 F_(1)offspring lines were all male,thus revealing a super-male status for their male parents. 360 F_(1) families contained male plants and female plants,proving that their male parents were male. This result is coincident with that of STS molecular marker genotyping. Collectively,the STS bimolecular functional marker can accurately classify the sex status(female,male and super-male plants)of asparagus,which has implication in the all-male molecular marker assisted breeding of asparagus.