大豆种质资源遗传多样性研究
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摘要
大豆(Soybean)为豆科大豆属一年生草本植物,种子含有丰富的蛋白质,原产我国,至今已有5000年的种植史。1956、1979、1990年3次全国范围内共收集栽培大豆遗传资源23587份,占世界23%,收集到不同类型的野生、半野生大豆种质6000多份,占世界野生资源的90%以上,这些丰富的核心种质材料对拓宽大豆育种的遗传基础,为解决大豆栽培生产上抗病、抗虫等优良基因的缺乏具有重要的意义,也为解决当前大豆栽培品种遗传基础狭窄提供了丰富的核心种质材料,本研究基于不同来源的大豆群体,从东北栽培、东北野生和大豆核心种质库中选取具有代表性的300份材料,分别从形态学、蛋白质、等位酶及DNA水平对其遗传多样性进行研究,为解决当前栽培大豆的遗传基础狭窄,提高大豆的产量与品质提供理论依据。
1、对供试群体形态多样性分析结果显示,生育期变异系数最大为核心种质材料,变异系数值为0.17;株高变异系数最大为核心种质材料,变异系数值为0.437,蛋白质含量变异系数最大为野生材料,变异系数值为0.109;脂肪含量变异系数最大为野生材料,变异系数值为0.18:百粒重变异系数最大也为野生材料,变异系数值为1.308;不同来源群体的5个数量性状(蛋白质含量、脂肪含量、百粒重、生育期、株高)的方差分析都达到了极显著水平;不同来源群体比较,野生材料蛋白质含量变异系数、百粒重变异系数最大;脂肪含量变异系数最大为供试群体的脂肪含量变异系数,变异系数值为0.20;不同来源群体5个质量性状(花色、种皮色、茸毛色、结荚习性、粒形)相比较,栽培大豆品种的结荚习性以亚有限居多,核心种质结荚习性以有限居多,野生材料结荚习性以无限居多:栽培大豆品种种皮色以淡黄、黄色居多,野生大豆材料种皮色以褐色、深褐色居多,核心种质种皮色以黄色居多,其它种皮色有之;栽培大豆籽粒形状以园粒、扁圆居多,核心种质籽粒形状以扁圆形、扁椭圆形居多,野生大豆粒形以扁椭圆居多。
2、对供试群体过氧化物酶等位酶谱带分析表明,过氧化物酶等位酶(PER),共检测到3个基因位点,15条酶带,所检测到的基因位点均为多态性位点,多态性位点的百分数达100%。过氧化物酶等位酶3个基因位点PER1、PER2和PER3,分别含有3条、5条和7条酶带。
3、对供试群体蛋白亚基的谱带分析表明,栽培大豆7S亚基含量变化范围11.93%～41.33%,平均值26.06%,方差为4.77,变异系数为18.3,其中含量高于平均值的品种有57个:野生大豆7S亚基含量变化范围19.19%～42.53%,均值为30.68%,方差为5.7,变异系数为18.6,其中含量高于平均值的种质有31个,栽培大豆11S亚基含量变化范围69.5%～40.33%,平均值60.27%,方差为4.27,变异系数为7.03,其中含量高于平均值的品种有55个;野生大豆11S亚基含量变化范围51.58%～71.47%,均值为63.94%,方差为3.75,变异系数为5.88,其中含量高于平均值的有种质30个:供试栽培和野生材料的11S/7S比值最大值和最小值分别为5.34和1.03;3.65和1.31,平均值为2.41:2.34,栽培大豆中11S/7S比值高于均值有41个,野生大豆种质比值高于平均值的有21个;供试群体不同来源间11S和7S球蛋白各组成亚基的含量差异显著,并且,不同来源种质11S球蛋白含量都高于7S球蛋白含量,大豆球蛋白11S/7S值具有较大的变异范围。
4、用经过筛选的50个SSR引物,对供试群体进行SSR检测,结果表明:50个引物共检测到742个位点,其中多态位点627个,多态位点高达84%,通过(UPGMA)聚类可以把供试群体分为4类,第一大类,包含167份材料,占供试材料总数的58%。从来源上看,大部分为栽培品种和核心种质;第2大类,包含71份材料,占供试材料总数的25%,包括部分栽培品种、核心种质和野生材料,第3类,包括43份材料,占供试材料总数的15%,全部为野生材料:第4类,包括7份材料,其中2份为栽培品种,5份为核心种质。
Soybean originated from China and belonged to Legume Glycine, which was Annual herbaceous plant. It called "Shu" in ancient China and it seed contained rich protein. Chinese people had grown soybean for five thousand years. Now it was generally grown nationwide, especially in Yangtze river basin and southwest, The optimal quality soybean was in northeast region. Soybean which was grown in others country directly or indirectly spread out by china. Because of its high nutritional value, soybean was called "The king of beans", "Meat in the field", "Green milk", and so on. Germplasm resources was very rich in our country, but the narrow genetic basis of soybean varieties had induced the reducing of yield and Serious influenced and restricted the development of soybean industry. Therefore, this research selected 300 representative materials from wild and cultivated soybean varieties and germplasm banks in Heilongjiang province based on the different groups to study its genetic diversity respectively from the morphology, protein, allozyme and DNA level, in order to broaden the genetic basis and increase the yield and quality of soybean.
Results of morphological diversity analysis on testing varieties showed that differentiation coefficient of growth period of core germplasm materials was maximum and was 0.17; differentiation coefficient of plant height of core germplasm materials was maximum and was 0.437; differentiation coefficient of protein content of wild germplasm materials was maximum and was 0.109; differentiation coefficient of fat content of wild germplasm materials was maximum and was 0.18; differentiation coefficient of hundred grain weight of wild germplasm materials was maximum and was 1.308; variance analysis of five quantitative traits of different original varieties achieved a significant level; most podding habits of cultivated soybean was sublimited, most podding habits of core germplasm materials was limited, most podding habits of wild soybean was unlimited; most seed capsule color of cultivated soybean was yellow and primrose yellow, most seed capsule color of wild soybean was brown and puce, most seed capsule color of core germplasm materials was yellow; most seed shape of cultivated soybean was round and oblate, most seed shape of core germplasm materials was oblate and flat oval, most seed shape of wild soybean was flat oval.
Mass spectrum analysis on peroxidase allozyme of testing varieties showed that 3 genetic loci and 15 enzyme bands were detected, all of detected genetic loci were polymorphic loci. 3 genetic loci (PER1、PER2 and PER3) of peroxidase allozyme respectively contained 3, 5 and 7 enzyme bands.
Mass spectrum analysis on protein subunits of testing varieties showed that variation range of 7S subunit content of cultivated soybean varieties was from 11.93% to 41.33%, mean of it was 26.06%, variance of it was 4.77, differentiation coefficient of it was 18.3, content of 57 varieties was higher than mean; variation range of 7S subunit content of wild soybean varieties was from 19.19% to 42.53%, mean of it was 30.68%, variance of it was 5.7, differentiation coefficient of it was 18.6, content of 31 varieties was higher than mean; variation range of 11S subunit content of cultivated soybean varieties was from 69.5% to 40.33%, mean of it was 60.27%, variance of it was 4.27, differentiation coefficient of it was 7.03, content of 55 varieties was higher than mean; variation range of 11S subunit content of wild soybean varieties was from 51.58% to 71.47%, mean of it was 63.94%, variance of it was 3.75, differentiation coefficient of it was 2.88, content of 30 varieties was higher than mean; maximum and minimum of 11S/7S of cultivated soybean varieties was 5.34 and 1.03, maximum and minimum of 11S/7S of wild soybean varieties was 3.65 and 1.31,the mean was 2.41 and 2.34 respectively, 11S/7S of 41 cultivated soybean varieties was higher than mean, 11S/7S of 21 wild soybean varieties was higher than mean, the difference of 11S and 7S protein subunit content in different original varieties were significant, 11S globulin content was higher than 7S globulin content, 1 ls/7s value had large range of variation.
SSR detecting of testing varieties was carried on by 50 selected SSR primers, the results showed that 742 loci was detected by 50 random primers, 627 of them was polymorphic loci and the rate reached 84%. Testing materials could be divided into 4 kinds by clustering analysis, frist kind accounted for 58% and contained 167 varieties which mostly was cultivated soybean varieties and core germplasm materials; second kind accounted for 25% and contained 71 varieties which was cultivated soybean varieties and wild soybean varieties and core germplasm materials; third kind accounted for 15% and contained 43 varieties which entirely was wild soybean varieties; fourth kind contained 7varieties, 2 of them was cultivated soybean varieties, 5 of them was core germplasm materials.
Comprehensive analysis results fully showed that selected soybean germplasm resources had abundant genetic diversity.
1、对供试群体形态多样性分析结果显示,生育期变异系数最大为核心种质材料,变异系数值为0.17;株高变异系数最大为核心种质材料,变异系数值为0.437,蛋白质含量变异系数最大为野生材料,变异系数值为0.109;脂肪含量变异系数最大为野生材料,变异系数值为0.18:百粒重变异系数最大也为野生材料,变异系数值为1.308;不同来源群体的5个数量性状(蛋白质含量、脂肪含量、百粒重、生育期、株高)的方差分析都达到了极显著水平;不同来源群体比较,野生材料蛋白质含量变异系数、百粒重变异系数最大;脂肪含量变异系数最大为供试群体的脂肪含量变异系数,变异系数值为0.20;不同来源群体5个质量性状(花色、种皮色、茸毛色、结荚习性、粒形)相比较,栽培大豆品种的结荚习性以亚有限居多,核心种质结荚习性以有限居多,野生材料结荚习性以无限居多:栽培大豆品种种皮色以淡黄、黄色居多,野生大豆材料种皮色以褐色、深褐色居多,核心种质种皮色以黄色居多,其它种皮色有之;栽培大豆籽粒形状以园粒、扁圆居多,核心种质籽粒形状以扁圆形、扁椭圆形居多,野生大豆粒形以扁椭圆居多。
2、对供试群体过氧化物酶等位酶谱带分析表明,过氧化物酶等位酶(PER),共检测到3个基因位点,15条酶带,所检测到的基因位点均为多态性位点,多态性位点的百分数达100%。过氧化物酶等位酶3个基因位点PER1、PER2和PER3,分别含有3条、5条和7条酶带。
3、对供试群体蛋白亚基的谱带分析表明,栽培大豆7S亚基含量变化范围11.93%～41.33%,平均值26.06%,方差为4.77,变异系数为18.3,其中含量高于平均值的品种有57个:野生大豆7S亚基含量变化范围19.19%～42.53%,均值为30.68%,方差为5.7,变异系数为18.6,其中含量高于平均值的种质有31个,栽培大豆11S亚基含量变化范围69.5%～40.33%,平均值60.27%,方差为4.27,变异系数为7.03,其中含量高于平均值的品种有55个;野生大豆11S亚基含量变化范围51.58%～71.47%,均值为63.94%,方差为3.75,变异系数为5.88,其中含量高于平均值的有种质30个:供试栽培和野生材料的11S/7S比值最大值和最小值分别为5.34和1.03;3.65和1.31,平均值为2.41:2.34,栽培大豆中11S/7S比值高于均值有41个,野生大豆种质比值高于平均值的有21个;供试群体不同来源间11S和7S球蛋白各组成亚基的含量差异显著,并且,不同来源种质11S球蛋白含量都高于7S球蛋白含量,大豆球蛋白11S/7S值具有较大的变异范围。
4、用经过筛选的50个SSR引物,对供试群体进行SSR检测,结果表明:50个引物共检测到742个位点,其中多态位点627个,多态位点高达84%,通过(UPGMA)聚类可以把供试群体分为4类,第一大类,包含167份材料,占供试材料总数的58%。从来源上看,大部分为栽培品种和核心种质;第2大类,包含71份材料,占供试材料总数的25%,包括部分栽培品种、核心种质和野生材料,第3类,包括43份材料,占供试材料总数的15%,全部为野生材料:第4类,包括7份材料,其中2份为栽培品种,5份为核心种质。
Soybean originated from China and belonged to Legume Glycine, which was Annual herbaceous plant. It called "Shu" in ancient China and it seed contained rich protein. Chinese people had grown soybean for five thousand years. Now it was generally grown nationwide, especially in Yangtze river basin and southwest, The optimal quality soybean was in northeast region. Soybean which was grown in others country directly or indirectly spread out by china. Because of its high nutritional value, soybean was called "The king of beans", "Meat in the field", "Green milk", and so on. Germplasm resources was very rich in our country, but the narrow genetic basis of soybean varieties had induced the reducing of yield and Serious influenced and restricted the development of soybean industry. Therefore, this research selected 300 representative materials from wild and cultivated soybean varieties and germplasm banks in Heilongjiang province based on the different groups to study its genetic diversity respectively from the morphology, protein, allozyme and DNA level, in order to broaden the genetic basis and increase the yield and quality of soybean.
Results of morphological diversity analysis on testing varieties showed that differentiation coefficient of growth period of core germplasm materials was maximum and was 0.17; differentiation coefficient of plant height of core germplasm materials was maximum and was 0.437; differentiation coefficient of protein content of wild germplasm materials was maximum and was 0.109; differentiation coefficient of fat content of wild germplasm materials was maximum and was 0.18; differentiation coefficient of hundred grain weight of wild germplasm materials was maximum and was 1.308; variance analysis of five quantitative traits of different original varieties achieved a significant level; most podding habits of cultivated soybean was sublimited, most podding habits of core germplasm materials was limited, most podding habits of wild soybean was unlimited; most seed capsule color of cultivated soybean was yellow and primrose yellow, most seed capsule color of wild soybean was brown and puce, most seed capsule color of core germplasm materials was yellow; most seed shape of cultivated soybean was round and oblate, most seed shape of core germplasm materials was oblate and flat oval, most seed shape of wild soybean was flat oval.
Mass spectrum analysis on peroxidase allozyme of testing varieties showed that 3 genetic loci and 15 enzyme bands were detected, all of detected genetic loci were polymorphic loci. 3 genetic loci (PER1、PER2 and PER3) of peroxidase allozyme respectively contained 3, 5 and 7 enzyme bands.
Mass spectrum analysis on protein subunits of testing varieties showed that variation range of 7S subunit content of cultivated soybean varieties was from 11.93% to 41.33%, mean of it was 26.06%, variance of it was 4.77, differentiation coefficient of it was 18.3, content of 57 varieties was higher than mean; variation range of 7S subunit content of wild soybean varieties was from 19.19% to 42.53%, mean of it was 30.68%, variance of it was 5.7, differentiation coefficient of it was 18.6, content of 31 varieties was higher than mean; variation range of 11S subunit content of cultivated soybean varieties was from 69.5% to 40.33%, mean of it was 60.27%, variance of it was 4.27, differentiation coefficient of it was 7.03, content of 55 varieties was higher than mean; variation range of 11S subunit content of wild soybean varieties was from 51.58% to 71.47%, mean of it was 63.94%, variance of it was 3.75, differentiation coefficient of it was 2.88, content of 30 varieties was higher than mean; maximum and minimum of 11S/7S of cultivated soybean varieties was 5.34 and 1.03, maximum and minimum of 11S/7S of wild soybean varieties was 3.65 and 1.31,the mean was 2.41 and 2.34 respectively, 11S/7S of 41 cultivated soybean varieties was higher than mean, 11S/7S of 21 wild soybean varieties was higher than mean, the difference of 11S and 7S protein subunit content in different original varieties were significant, 11S globulin content was higher than 7S globulin content, 1 ls/7s value had large range of variation.
SSR detecting of testing varieties was carried on by 50 selected SSR primers, the results showed that 742 loci was detected by 50 random primers, 627 of them was polymorphic loci and the rate reached 84%. Testing materials could be divided into 4 kinds by clustering analysis, frist kind accounted for 58% and contained 167 varieties which mostly was cultivated soybean varieties and core germplasm materials; second kind accounted for 25% and contained 71 varieties which was cultivated soybean varieties and wild soybean varieties and core germplasm materials; third kind accounted for 15% and contained 43 varieties which entirely was wild soybean varieties; fourth kind contained 7varieties, 2 of them was cultivated soybean varieties, 5 of them was core germplasm materials.
Comprehensive analysis results fully showed that selected soybean germplasm resources had abundant genetic diversity.
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