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大豆种子成熟至收获期间如遇高温高湿天气,种子活力及活性会急剧下降,这就是所谓的田间老化(field weathering)。田间老化是热带、亚热带地区大豆生产的主要限制因素之一。本研究旨在寻找与田间老化性状相连锁的DNA标记并将其应用于辅助选择育种。为此,利用修改的培养箱老化法和人工控制老化法对大豆品种Chiangmai 60(敏感),GC10981(抵抗)及其F2群体(139个体)进行了鉴定。在两种处理条件下,F2代群体的种子发芽率及活性均为正态分布,说明大豆种子田间老化抗性受多个基因控制。根据F2代个体的种子发芽率及活性,6个高抗个体及7个高感个体的DNA分别被混合为抗性池和感性池,并利用AFLP标记进行了混合群体分析(BulkSegregant Analysis)。从扩增的2162个标记中,发现了5个可能于大豆种子田间抗性相连锁的片段。通过DNA克隆和测序,设计了5对引物用于从大豆总DNA中扩增相应的片段。其中3对引物扩增的片段差异太小或未能扩增正确大小的片段,没能用于F2群体。引物Eaag/Mcac-233和Eact/Mctt-157能扩增出差异明显的多态性,通过对F2代群体的分析,这2个标记属同一连锁群,遗传距离为25.8c M。QTL分析结果显示有一个QTL位于这两个标记之间,距Eaag/Mcac-233约14c M,可以解释29.7%的变异。用这两对引物对整个F2群体进行筛选,20个个体属于抗性群体,结合抗性鉴定的结果,7个个体被用于与Chiangmai60进行回交。18个BC1F1个体(41·9%)的抗性高于其亲本的平均值。说明这些标记进行可以被用于大豆田间老化抗性的辅助筛选研究。
Soybean seeds in the case of ripe to harvest in case of high temperature and humidity weather, seed vigor and activity will drop sharply, which is the so-called field weathering (field weathering). Field aging is one of the major constraints on soybean production in tropical and subtropical regions. The aim of this study was to find DNA markers linked to field-aged traits and to apply them to assisted selection breeding. To this end, soybean cultivars Chiangmai 60 (susceptible), GC10981 (resistant) and their F2 population (139 individuals) were identified using a modified incubator and artificial aging method. Under the two treatment conditions, the seed germination rate and activity of F2 population were both normal distribution, indicating that the aging resistance of soybean seed was controlled by multiple genes. According to the seed germination rate and activity of F2 individuals, DNAs from 6 highly resistant individuals and 7 highly susceptible individuals were mixed into resistant and susceptible pools respectively, and the Bulk Segregant Analysis was performed using AFLP markers. From the 2162 markers amplified, five fragments that could be linked to resistance in the soybean seed were found. Through DNA cloning and sequencing, five pairs of primers were designed to amplify the corresponding fragments from total soybean DNA. Three pairs of primers amplified fragments that were too small or failed to amplify the correct size of the fragment, failing to be used in the F2 population. The primers Eaag / Mcac-233 and Eact / Mctt-157 can amplify the significantly different polymorphism. By analyzing F2 population, the two markers belong to the same linkage group with a genetic distance of 25.8cM. QTL analysis showed that one QTL was located between these two markers, about 14cM away from Eaag / Mcac-233, accounting for 29.7% of the variance. The entire F2 population was screened with these two pairs of primers, and 20 individuals belonged to the resistant population. Seven individuals were used for backcrossing with Chiangmai60 in combination with the results of resistance identification. The resistance of 18 BC1F1 individuals (41.9%) was higher than the average of their parents. These markers are described as a secondary screening study that can be used for aging resistance in soybean fields.