水稻第2号染色体深根比主效QTL-qRDR-2的精细定位

发布时间：2018-11-13 09:53

【摘要】：水稻(Oryza sativa L.)是世界上重要的粮食作物之一,世界上有一半的人口以稻米为食。干旱是影响产量的重要非生物胁迫之一。近些年,我国城镇化进程的加快,更是加剧了我国水资源的短缺,因此,研究节水抗旱稻至关重要。根系是水稻重要的吸水器官,较深的根系分布层可以增加水稻的吸水量,提高水稻的避旱性。对控制深根比基因的精细定位,能够帮助我们揭示抗旱机理,加快节水抗旱稻选育进程,同时对保障我国粮食安全也具有重要意义。本实验室自2004年以来利用珍汕97B与IRAT109构建重组自交系(Recombinant Inbred Lines,RIL)群体,并利用213个SSR分子标记构建了覆盖全基因组的图谱。通过多年多点鉴定,共检测到四个深根比主效QTL,分别位于第1、2、4、7号染色体。通过回交,结合目标QTL连锁标记前景选择和全基因组背景选择,构建以珍汕97B为背景,含有目标QTL的4个近等基因系(Near Isogenic Line,NIL)。在此基础上,本文针对位于第2号染色体上深根比主效QTL-q RDR-2进行了精细定位,结果如下:1.BC_4F_1前景与背景检测:本实验室于2015年春,完成含有目标QTL-q RDR-2的BC_4F_1构建,结合前景连锁标记及背景标记基因型检测,选择目标区段连锁标记基因型双杂合,背景与珍汕97B尽量相似单株,在63个BC_4F_1中共筛选出32个单株。2.q RDR-2重组交换单株的筛选及重组交换位置的确定:将含有目的片段的近等基因系BC_4F_1发展为含有8000株的BC_4F_2群体,利用初定位区间两侧连锁标记RM6与RM240进行重组交换单株的筛选,共筛选出289株重组交换单株。通过分析重组交换单株目标区段标记基因型,发现其中14个标记发生重组,包括22个重组交换单株。3.q RDR-2精细定位:本课题于2016年6月,将22个重组交换单株的自交后代种植于上海金山廊下基地,每个重组交换单株种植42个BC_4F_3单株。利用“篮子法”对每个单株进行表型鉴定,并结合基因型检测,将QTL-q RDR-2定位于分子标记Q2-323与Q2-12之间,物理距离为570kb。4.第二次重组交换单株的筛选及重组交换位置的确定:为了进一步缩小区间距离,在第一次定位的基础上,利用QTL区间两侧连锁标记Q2-323与Q2-12,从4000个F_4单株中,筛选出106个重组交换单株。通过分析重组交换单株目标区段标记基因型,发现其中6个标记发生重组,共包括12个重组交换单株。5.候选基因预测:通过生物信息学分析,并结合本实验室根尖RNA-seq数据,在目标区段预7个候选基因:LOC_Os02g48360、LOC_Os02g48710、LOC_Os02g48770、LOC_Os02g49160、LOC_Os02g49440、LOC_Os02g49460、LOC_Os02g49720,其中LOC_OS02g49160为已知基因OsPIN1。
[Abstract]:Rice (Oryza sativa L.) It is one of the most important food crops in the world. Half of the world's population feeds on rice. Drought is one of the important abiotic stresses affecting yield. In recent years, the quickening process of urbanization in China has aggravated the shortage of water resources in China, so it is very important to study water-saving and drought-resistant rice. Root system is an important water absorption organ of rice. The deeper root distribution layer can increase the water absorption of rice and improve the drought avoidance of rice. The precise mapping of the genes controlling the deep root ratio can help us to reveal the mechanism of drought resistance, accelerate the breeding process of water-saving drought-resistant rice, and also play an important role in ensuring the food security in China. Since 2004, Zhenshan 97B and IRAT109 were used to construct recombinant inbred line (Recombinant Inbred Lines,RIL) population, and 213 SSR molecular markers were used to construct the genome map. After many years of identification, four deep root specific dominant QTL, were found to be located on chromosome 1, chromosome 2 and chromosome 7, respectively. By backcrossing, combining target QTL linkage marker foreground selection and whole genome background selection, four near-isogenic lines (Near Isogenic Line,NIL with Zhenshan 97B background and target QTL were constructed. On this basis, a detailed mapping of deep root specific dominant QTL-q RDR-2 on chromosome 2 was carried out. The results are as follows: 1.BC_4F_1 foreground and background test. The construction of BC_4F_1 containing target QTL-q RDR-2 was completed. Combined with foreground linkage marker and background marker genotype detection, the target segment linkage marker genotype double heterozygosity was selected, and the background was similar to that of Zhenshan 97B single plant. A total of 32 individual plants were screened out of 63 BC_4F_1. 2. Screening of Q RDR-2 recombinant exchange single plants and determination of the recombination exchange site: BC_4F_1, a near-isogenic line containing the target fragment, was developed into a BC_4F_1 containing 8000 strains. The BC_4F_2 community, Two sides linkage marker RM6 and RM240 were used to screen the recombinant exchange single plant, and a total of 289 recombinant exchange single plants were screened. Based on the analysis of target segment marker genotypes of recombinant exchange single plant, it was found that 14 of these markers were recombined, including 22 recombinant exchange single plants. 3. Q RDR-2 fine location: this study was conducted in June, 2016. The inbred progenies of 22 recombinant exchange plants were planted in Jinshan Corridor, Shanghai, and 42 BC_4F_3 plants were planted per recombination exchange single plant. The QTL-q RDR-2 was located between Q2-323 and Q2-12 with the physical distance of 570kb.4. the phenotype of each plant was identified by "basket method" and combined with genotype detection. Screening of the second recombination exchange single plant and determination of the recombination exchange location: in order to further reduce the interval distance, on the basis of the first location, the linkage markers Q2-323 and Q2-12 of the two sides of the QTL interval were used, and the sequence of Q2-323 and Q2-12 were obtained from 4 000 FSCL plants. 106 recombinant exchange single plants were screened. By analyzing the target segment marker genotypes of the recombinant exchange single plant, it was found that 6 of the markers were recombined, including 12 recombinant exchange single plants. Candidate gene prediction: through bioinformatics analysis, combining with RNA-seq data of root tip of our laboratory, 7 candidate genes: LOC_Os02g48360,LOC_Os02g48710,LOC_Os02g48770,LOC_Os02g49160,LOC_Os02g49440,LOC_Os02g49460,LOC_Os02g49720, in target region are predefined. Where LOC_OS02g49160 is a known gene OsPIN1.
【学位授予单位】：华中农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S511

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