大豆(Glycine max(L.)Merr.)致敏蛋白Gly m Bd 30K检测方法及低致敏优异种质鉴定  

Detection Method of Allergen Gly m Bd 30K and Identification of Elite Accessions with Low Allergenicity in Soybean(Glycine max(L.)Merr.)

作　　者：周明明 张明俊 王俊[1] 邱丽娟[2] ZHOU Ming-ming;ZHANG Ming-jun;WANG Jun;QIU Li-juan(Yangtze University,College of Agriculture,Hubei Jingzhou 434025;National Key Facility for Crop Gene Resources and Genetic Improvement/Key Laboratory of Soybean Biology in Beijing(MOA)/Institute of Crop Sciences,Chinese Academy of Agricultural Sciences,Beijing 100081)

机构地区：[1]长江大学农学院,湖北荆州434025 [2]农作物基因资源与遗传改良国家重大科学工程/农业部北京大豆生物学重点实验室/中国农业科学院作物科学研究所,北京100081

出　　处：《植物遗传资源学报》2021年第1期149-156,共8页Journal of Plant Genetic Resources

基　　金：农作物种质资源保护与利用专项(2019NWB036-05);中国农科院科技创新工程项目。

摘　　要：Gly m Bd 30K蛋白是大豆中主要的免疫显性过敏原之一,会引起人和牲畜腹泻和肠道炎症等过敏反应。因此,发掘低Gly m Bd 30K蛋白含量优异种质对于培育优质大豆品种具有重要意义。为了获得致敏蛋白Gly m Bd 30K低含量的优异种质,根据Gly m Bd 30K蛋白的190-379aa多肽序列制备多克隆抗体;对来源于山西省的29份种质,利用Western blot技术对其Gly m Bd 30K蛋白含量进行检测;并扩增和分析Gly m Bd 30K基因序列;利用荧光定量PCR技术对筛选出的低Gly m Bd 30K含量种质进行表达分析。结果表明:从参试种质中鉴定出2份Gly m Bd 30K蛋白含量低的种质,Gly m Bd 30K蛋白低含量种质的鉴定效率为6.9%,分别是来自太原市的大豆种质134(ZDD02046)和大青豆(ZDD02174),其Gly m Bd 30K基因序列与Willams82相比,在启动子上都有TA重复序列变异,134(ZDD02046)有8次TA重复,大青豆(ZDD02174)有42次TA重复,表达分析结果显示,134(ZDD02046)的转录水平显著低于大青豆(ZDD02174),推测启动子上TA多态性可能影响了其转录水平。本研究建立Gly m Bd 30K蛋白含量测定的方法,鉴定出2份低蛋白含量的优异种质,为今后选育优质蛋白组合的大豆新品种提供了技术和材料支撑。Gly m Bd 30 K protein is recognized as a major immunodominant allergen in soybean,and this protein can lead to anaphylaxis such as diarrhea and intestinal inflammation in humans and livestock.The exploration of germplasms with lower Gly m Bd 30 K content is important in breeding for elite functional soybean varieties.In this study,we generated the polyclonal antibody targeting to the 190-379 aa peptides of the Gly m Bd 30 K protein,followed by quantifying the Gly m Bd 30 K protein using Western blot in 30 soybean accessions that were derived from Shanxi province.The genomic sequence of the Gly m Bd 30 K gene and its transcriptional level was further conducted.As a result,two germplasms(134,ZDD02046,Daqingdou,ZDD02174)with lower Gly m Bd 30 K protein content have been identified,the identification efficiency is 6.9%.Both accessions displayed 8 and 42 TA repeats respectively in the promoter of the Gly m Bd 30 K gene if compared with that of Willams82.The transcription level in 134(ZDD02046)was found to be significantly lower than that of Daqingdou(ZDD02174).It is speculated that TA polymorphism in the promoter may associate with the Gly m Bd 30 K expression.Collectively,this study established a strategy on the quantification of Gly m Bd 30 K protein content and identified two accessions with lower Gly m Bd 30 K protein,which will provide technical and material supports in breeding for new soybean varieties with high-quality proteins.

关 键 词：大豆 过敏原 Gly m Bd 30K 种质筛选 Western blot 

分 类 号：S565.1[农业科学—作物学]