| Brassica napus L.has the characteristics of tall plant architecture,stable yield and high oil content in seeds,so it is widely planted in China as the main oil crop.The traditional Brassica napus L.planting method takes time and effort to seriously limit the enthusiasm of workers.Therefore,improving the plant architecture of Brassica napus L.to cultivate new varieties suitable for mechanized planting is beneficial to improve its production efficiency.The main stem of dicotyledonous plants has the functions of supporting organs,transporting nutrients and storing substances,which plays a key role in the process of plant architecture formation and yield formation.At present,most varieties of Brassica napus are single main stem,and its yield per plant is nearly 1/4 from the pods of the main inflorescence,so increasing the number of main stems is of great significance to improve the yield of Brassica napus.In this study,a multiple main stem mutant 90-12 produced by crossing between Brassica napus L.and Brassica campestris L.was used as material.Through field identification,it was inferred that the multiple main stem phenotype was controlled by a single recessive nuclear locus.Combined with BSA-seq technology,the genetic linkage map was constructed,and the markers linked to the multiple main stem phenotype were screened to provide a candidate interval for fine mapping to control the related loci of 90-12 multiple main stem traits.The main results are as follows:1.Based on the investigation and statistics of agronomic characters of 90-12 and Zhongshuangll(ZS11),it was found that the number of main stems of 90-12 was significantly more than that of ZS11,and the number of branches,the number of leaves at the end of flowering and the height of the first branch of 90-12 increased significantly by 54%,47%and 18%compared with ZS11.There was no significant difference in 1000grain weight and oil content between 90-12 and ZS11.Through the planting investigation in spring/winter rape area(Badong/Wuhan)for three consecutive years,it was found that the phenotype of 90-12 main stems was always stable and was not regulated by environmental factors such as light and temperature.2.The seedling development process of 90-12 and ZS11 was investigated,and it was found that there was no significant difference in seedling morphology within 10 days after sowing.After 15 days of sowing,the increasing rate of leaf number of 90-12 was higher than that of ZS11,and the difference gradually expanded with the growth and development of seedlings.The cytological observation of SAM in different stages of seedlings showed that the tip of 90-12 was irregular and produced multiple leaf primordia on the 20th day,while the top of ZS11 was hemispherical and smooth.On the 30th day,new axillary bud primordia were produced at the angle between the leaf primordia of 90-12 SAM and young leaves,while ZS11 did not produce excess axillary bud primordia.3.The stem slices of 90-12 and ZS11 were observed and analyzed,and it was found that the cortex of ZS11 was thinner and the cells arranged neatly and compactly,while that of 90-12 was thicker,the arrangement of cells was loose,and there was a region of cell loss.Statistics showed that the area of Middle columnar cell and the number of vascular bundles in 90-12 decreased significantly by 44%and 40%respectively compared with ZS11,and the lignin and cellulose contents in 90-12 decreased significantly by 23%and 7%respectively compared with ZS 11.4.Based on the field investigation and genetic analysis of F1 single plant and F2 population obtained from 90-12 ZS11,it was found that F1 single plant was single main stem phenotype,F2 population had 320 single main stem single plant and 96 multiple main stem single plant.After χ2 test(χ2=0.82,p=0.37),the segregation ratio was consistent with 3:1,so the multiple main stem phenotype was controlled by a single recessive locus,which was named Mms(Multiple main stem).5.In F2 population,30 plants of extreme single main stem and 30 plants of extreme multiple main stem were selected to construct N-Bulk and M-Bulk respectively.N/M-Bulk was analyzed by BSA-seq(Bulked Segregant Analysis),and a total of 621563 InDel loci were obtained.Then 632 InDel markers were developed and polyacrylamide gel electrophoresis(Polyacrylamide Gel Electrophoresis,PAGE)was used to analyze the linkage of F2 population to construct a genetic linkage map with a total length of 1413.2 cM.The map is divided into 21 linkage groups with a total of 222 markers.The number of markers in each linkage group varies from 6 to 21,and the average distance between adjacent markers is distributed between 2.4 and 11.3 cM,which is used for the preliminary mapping of multiple main stem gene Mms.6.According to the results of genetic linkage map and BSA-seq,it is speculated that there are related loci of multiple main stem gene Mms in the interval of A09 chromosome 14.01-51.03 Mb.The 52 InDel markers were developed in this interval and preliminary linkage analysis was conducted in F3 population composed of extremely single/multiple main stem.A marker ChrA9-14140308 linked to multiple main stem phenotype was screened.The 236 markers were further developed in the interval of A09 chromosome 10.00-40.00 Mb and linkage analysis was carried out using 78 plants of extremely single main stem and 127 plants of extremely multiple main stem(F3).As a result,the multiple main stem gene Mms was initially located in the interval of A09 chromosome 20.99-39.11 Mb. |
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