水稻qPC1基因启动子功能性变异位点的鉴定及其转运功能  

Identification of functional mutation sites of qPC1 gene promoter in rice and analysis of its transport function

作　　者：彭波 邱静 彭娟 何璐璐 孔冬艳 孙晓宇 刘岩 阿新祥[3] 靳可心 孙艳芳 庞瑞华 周伟 赵金会 汪全秀 PENG Bo;QIU Jing;PENG Juan;HE Lu-lu;KONG Dong-yan;SUN Xiao-yu;LIU Yan;A Xin-xiang;JIN Ke-xin;SUN Yan-fang;PANG Rui-hua;ZHOU Wei;ZHAO Jin-hui;WANG Quan-xiu(School of Life Sciences,Xinyang Normal University/Dabie Mountain Agricultural Biological Resource Protection and Utilization Re-search Institute,Xinyang,Henan 464000,China;Xinyang Plant Protection and Inspection Station,Xinyang,Henan 464000,China;Institute of Biotechnology and Germplasm Resources,Yunnan Academy of Agricultural Sciences/Yunnan Rice Seed Resource Observation Experimental Station,Ministry of Agriculture,Kunming 650223,China)

机构地区：[1]信阳师范大学生命科学学院/大别山农业生物资源保护与利用研究院,河南信阳464000 [2]信阳市植保植检站,河南信阳464000 [3]云南省农业科学院生物技术与种质资源研究所/农业部云南稻种资源观测实验站,昆明650223

出　　处：《西南农业学报》2023年第12期2573-2582,共10页Southwest China Journal of Agricultural Sciences

基　　金：国家自然科学基金项目(U2004141,U1604110,31801332);河南省高等学校青年骨干教师培养计划项目(2019GGJS162);河南省科技攻关项目(222102110141,222102110115,212102110249);信阳市创新专项(20210007);信阳师范学院2022年研究生科研创新基金项目(2022KYJJ066,2022KYJJ068)。

摘　　要：【目的】鉴定水稻种子中蛋白质含量的正调控基因qPC1启动子的功能性变异位点,开发qPC1基因分子标记,并解析其转运功能,为阐明qPC1基因的生物学功能及其利用qPC1基因进行分子育种提供理论依据。【方法】采用基因定点突变、水稻原生质体瞬时转化技术和双荧光素酶报告法鉴定qPC1基因启动子功能性变异位点,利用缺陷型酵母互补试验以及水稻根部氨基酸的吸收试验解析qPC1的转运功能。【结果】相对于南洋占qPC1序列,珍汕97 qPC1启动子-7~-12 bp位置处6个碱基(CACAGA)的缺失将导致其启动子活性显著增加。对此设计对应的功能性分子标记PB13,并利用PCR技术在珍汕97与南洋占的重组自交系群体中进行验证。在缺陷型酵母互补试验中发现qPC1蛋白能够转运γ-氨基丁酸、瓜氨酸、天冬氨酸、脯氨酸、精氨酸和谷氨酸等多种氨基酸,且发现qPC1能促进水稻根对多种氨基酸的吸收,并对苏氨酸、丝氨酸、甘氨酸、丙氨酸、脯氨酸、谷氨酸和酸性氨基酸具有较快的吸收与转运速率。【结论】水稻qPC1基因启动子区-7~-12 bp位置为其功能性变异位点,qPC1在酵母中能够参与多种氨基酸转运,且在水稻根中能促进氨基酸的吸收与转运,即qPC1蛋白可能是一种广谱性氨基酸转运蛋白。该试验结果为进一步全面阐明qPC1基因调控水稻种子中蛋白质含量的分子机制并利用qPC1基因进行优质水稻新品种的选育提供了重要信息。【Objective】Identifying the functional variation sites of the qPC1 promoter,a positive regulatory gene for grain protein content in rice,developing corresponding molecular markers,and analyzing the transport function of the qPC1 will provide a theoretical basis for eluci⁃dating the biological function of the qPC1 gene and its use in molecular breeding.【Method】The functional variation sites of qPC1 gene pro⁃moter were identified by site directed mutagenesis,transient transformation technology of rice protoplasts and double luciferase reporting Southwest China Journal of Agricultural Sciences method.The transport function of qPC1 was analyzed by deficient yeast complementation test and the absorption experiment of amino acids in rice roots.【Result】Compared to the Nanyangzhan qPC1 sequence,the deletion of a 6⁃bp(CACAGA)at the-7 bp to-12 bp position in the Zhenshan97 qPC1 promoter would significantly increase the activity of its qPC1 promoter.Based on the above characteristics,a corre⁃sponding functional molecular marker PB13 was designed and validated using PCR technology in the recombinant inbred line population of Zhenshan97 and Nanyangzhan.Through the complementary experiment of defective yeast,it was found that qPC1 protein could be transpor⁃ted various amino acids such asγ⁃aminobutyric acid,citrulline,aspartic acid,proline,arginine and glutamic acid.Through the amino acid absorption test of rice roots,it was found that the qPC1 could promote the absorption of various amino acids by rice roots,and had a fast ab⁃sorption and transport rate for threonine,serine,glycine,alanine,proline,glutamic acid and acidic amino acids.【Conclusion】The promot⁃er region of the qPC1 in rice,located at-7 bp to-12 bp,was its functional variation site.The qPC1 could participate in the transport of various amino acids in yeast and promote the absorption and transport of amino acids in rice roots.Therefore,the qPC1 may be a broad⁃spec⁃trum amino acid transport protein.The results of this experiment pr

关 键 词：功能性位点 qPC1基因 转运功能 启动子 

分 类 号：S511.01[农业科学—作物学]