旱稻IRAT109辐射突变体的培育与类病斑基因的初步定位
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摘要
作为一份典型的旱稻品种,IRAT109被广泛用于抗旱生理研究、基因/QTL作图和节水抗旱稻的遗传改良。水稻功能基因研究最直接最有的方法是构建饱和的基因突变群体。本研究以该品种为核心材料,进行辐射诱变,建立功能基因发掘的研究体系。
本研究分为两个部分。第一部分是260份突变体的繁育加代,包括背景分析、性状调查、突变类型划分归类、群体构建等。第二部分是选择一个旱敏感、叶片、叶鞘上有类病斑的突变体群体,对该突变基因进行初步定位。主要结果如下:
1.用47对常用于水稻品种鉴定的SSR标记,对260份突变体进行背景鉴定,结合标记的统计结果与田间性状调查结果,共剔除非IRAT109来源的材料51份。调查209份IRAT109来源突变体材料的性状,根据表型,按照叶、茎、穗型、颖壳、抗旱性等性状突变,分为20多个类型。用性状变化明显、能够稳定遗传的突变类型构建群体,共构建142个群体,用于对突变基因的分析。对于从209份突变体中筛选出34份对干旱胁迫有不同抗性的材料,以抗旱指数为基础,综合考虑各突变体的农艺性状表现,经大棚抗旱鉴定,初步鉴定了34份突变体材料的抗旱能力。其中有6份材料抗旱性明显减弱,表现为旱敏感。
2.调查发现,在类病斑突变体的两个F_2分离群体中,正常植株与类病斑植株的分离比例分别为:12.58:1和14.60:1,卡方检验表明,两种分离都符合15:1的比例。这表明类病斑性状可能同时受两对隐性基因控制。以9311×07TB-157(类病斑突变体lesion mimic mutant)的F2群体为定位群体,用分布于12条染色体上的1152对SSR标记,检测9311,07TB-157基因组之间的扩增差异,用有差异的引物分析其在F_2群体中隐性单株的分离情况,最终将突变基因定位在第10号染色体上的RM1374和RM4771附近,遗传连锁分析表明,类病斑突变体的突变基因lesion mimicinitiation(lmi)与RM1374的遗传距离为22.67cM,另一个位点与RM4771的遗传距离为18.62cM。RM1374与RM4771之间的物理距离为1.44Mb,相对较远,初步认为RM1374与RM4771可能分别与导致出现类病斑性状的两个基因连锁。
As one of typical upland rice variety,IRAT109 is used widely for physiological research of drought resistance,gene/QTLmapping,genetic improvement of water saving and drought resistance rice.The most direct and available research method for functional gene in rice is to construct a saturated mutant gene library.This study made IRAT109 as the core material;construct a radiation induction mutant library,set up the research system of functional gene in rice.
This study consists of two parts.The first part is the cultivation of 260 Radiation-induced mutants of upland rice IRAT109.This part includes background analysis,traits investigation,classification,construction of populations of 260 mutants and so on.Part three is gene mapping of one of the 260 mutant.The mutant that be chosen is susceptible to drought stress and have lesion mimic on leaves and sheath,that is a distinguished trait.The main results were as follows:
1.47 SSR makers are selected for the background detection of 260 mutants. Comprehensive analysis the result of SSR makers and agronomic characters,we identify that 51 mutants are not the self-pollinated progenies of radiation-induced mutants.After eliminating 51 lines,we still have 209 mutants which come from original parent IRAT109. Survey the traits of 209 mutant,we classify them into 20 more type according to the phenotype,such as leaf mutant,stem mutant,panicle type mutant,rice glume mutant, drought resistance mutant and so on.Some mutant whose character variation significantly and has genetic stability are used for constructing population,finally we get 142 populations that can be used for analysis mutant gene.34 mutants whose drought resistance ability is at various levels are selected from 209 mutants.Basing on drought-resistance coefficient and comprehensive agronomic characters,the drought resistance levels of 34 mutants are identified initially,the drought resistance of 6 mutants is decreased obviously.They are divided into drought sensitive type.
2.The field investigation show that the ratio of normal plant and lesion mimic initiation plant in two F_2 population of lesion mimic mutant is 12.58:1 and 14.60:1,the chi-square test show that they are fit the ratio of 15:1.So the lesion mimic trait might controlled by two recessive genes.We use F_2 population which the filial generation of 9311 and 07TB-157(lmi mutant) as the mapping population.To detect the genomic differences between two parents,1152 SSR makers were selected.The makers which show polymorphism between two parents are used for analyzing all the recessive individual plant.The gene loci controlling lmi were tagged by RM1374 and RM4771 on the chromosome 10.Genetic linkage analysis show that the genetic distance between one of lmi loci and RM1374 is 22.67cM,and the genetic distance between another lmi loci and RM4771 is 18.62cM.Base on the sequence data of Nipponbare,the physical distance between RM1374 and RM4771 is 1.44Mb,RM1374 and RM4771 may be linked with the two genes which control the lesion mimic trait.
本研究分为两个部分。第一部分是260份突变体的繁育加代,包括背景分析、性状调查、突变类型划分归类、群体构建等。第二部分是选择一个旱敏感、叶片、叶鞘上有类病斑的突变体群体,对该突变基因进行初步定位。主要结果如下:
1.用47对常用于水稻品种鉴定的SSR标记,对260份突变体进行背景鉴定,结合标记的统计结果与田间性状调查结果,共剔除非IRAT109来源的材料51份。调查209份IRAT109来源突变体材料的性状,根据表型,按照叶、茎、穗型、颖壳、抗旱性等性状突变,分为20多个类型。用性状变化明显、能够稳定遗传的突变类型构建群体,共构建142个群体,用于对突变基因的分析。对于从209份突变体中筛选出34份对干旱胁迫有不同抗性的材料,以抗旱指数为基础,综合考虑各突变体的农艺性状表现,经大棚抗旱鉴定,初步鉴定了34份突变体材料的抗旱能力。其中有6份材料抗旱性明显减弱,表现为旱敏感。
2.调查发现,在类病斑突变体的两个F_2分离群体中,正常植株与类病斑植株的分离比例分别为:12.58:1和14.60:1,卡方检验表明,两种分离都符合15:1的比例。这表明类病斑性状可能同时受两对隐性基因控制。以9311×07TB-157(类病斑突变体lesion mimic mutant)的F2群体为定位群体,用分布于12条染色体上的1152对SSR标记,检测9311,07TB-157基因组之间的扩增差异,用有差异的引物分析其在F_2群体中隐性单株的分离情况,最终将突变基因定位在第10号染色体上的RM1374和RM4771附近,遗传连锁分析表明,类病斑突变体的突变基因lesion mimicinitiation(lmi)与RM1374的遗传距离为22.67cM,另一个位点与RM4771的遗传距离为18.62cM。RM1374与RM4771之间的物理距离为1.44Mb,相对较远,初步认为RM1374与RM4771可能分别与导致出现类病斑性状的两个基因连锁。
As one of typical upland rice variety,IRAT109 is used widely for physiological research of drought resistance,gene/QTLmapping,genetic improvement of water saving and drought resistance rice.The most direct and available research method for functional gene in rice is to construct a saturated mutant gene library.This study made IRAT109 as the core material;construct a radiation induction mutant library,set up the research system of functional gene in rice.
This study consists of two parts.The first part is the cultivation of 260 Radiation-induced mutants of upland rice IRAT109.This part includes background analysis,traits investigation,classification,construction of populations of 260 mutants and so on.Part three is gene mapping of one of the 260 mutant.The mutant that be chosen is susceptible to drought stress and have lesion mimic on leaves and sheath,that is a distinguished trait.The main results were as follows:
1.47 SSR makers are selected for the background detection of 260 mutants. Comprehensive analysis the result of SSR makers and agronomic characters,we identify that 51 mutants are not the self-pollinated progenies of radiation-induced mutants.After eliminating 51 lines,we still have 209 mutants which come from original parent IRAT109. Survey the traits of 209 mutant,we classify them into 20 more type according to the phenotype,such as leaf mutant,stem mutant,panicle type mutant,rice glume mutant, drought resistance mutant and so on.Some mutant whose character variation significantly and has genetic stability are used for constructing population,finally we get 142 populations that can be used for analysis mutant gene.34 mutants whose drought resistance ability is at various levels are selected from 209 mutants.Basing on drought-resistance coefficient and comprehensive agronomic characters,the drought resistance levels of 34 mutants are identified initially,the drought resistance of 6 mutants is decreased obviously.They are divided into drought sensitive type.
2.The field investigation show that the ratio of normal plant and lesion mimic initiation plant in two F_2 population of lesion mimic mutant is 12.58:1 and 14.60:1,the chi-square test show that they are fit the ratio of 15:1.So the lesion mimic trait might controlled by two recessive genes.We use F_2 population which the filial generation of 9311 and 07TB-157(lmi mutant) as the mapping population.To detect the genomic differences between two parents,1152 SSR makers were selected.The makers which show polymorphism between two parents are used for analyzing all the recessive individual plant.The gene loci controlling lmi were tagged by RM1374 and RM4771 on the chromosome 10.Genetic linkage analysis show that the genetic distance between one of lmi loci and RM1374 is 22.67cM,and the genetic distance between another lmi loci and RM4771 is 18.62cM.Base on the sequence data of Nipponbare,the physical distance between RM1374 and RM4771 is 1.44Mb,RM1374 and RM4771 may be linked with the two genes which control the lesion mimic trait.
引文
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