| Chinese cabbage?Brassica rapa L.ssp.Pekinensis,AA?is one of the most important vegetable crops in China,belonging to Brassica rapa species of Brassica genus in Cruciferae family.Wide variation exists in Chinese cabbage germplasm,with various ecological cultivar groups.Most of agronomic traits including leaf and heading development are quantity traits and have complex genetic basis.It is necessary to explore the regulated mechanism at molecular levels.In recent years,few researches about the identification of some candidate genes related to leaf and heading in Brassica rapa were published,however it is not very clear about key genes and their regulations.The genome sequence of Chinese cabbage chiifu401 indicated that the genes related to auxin maybe one of important gene families affecting morphological organ development.Above information offers help in mapping genes and researching expression pattern for leaf and heading development in Brassica rapa.In this research,179 Chinese cabbage accessions were used to study the variation in morphological traits and to perform association mapping.And the doubled haploid DH88and F2 populations derived from parents of Pak choi and heading Chinese cabbage were used to as materials,combing with of constructing high density genetic maps and phenotyping leaf and heading related traits,QTL analysis for target traits were performed.Furthermore,nine Chinese cabbage genotypes representing different heading types?folding,overlapped,bundling center and non-head?and characteristics?early,middle and late?were used to identify expression patterns at eight development stages of 10 candidate genes related to heading and auxin.The purpose is to explore the genetic variation of important agronomic traits including leaf and heading development in the used populations,and to detect main regulating chromosome regions and fine related QTLs.The co-localized genes and molecular markers related to QTL will be checked and regulated patterns of candidate genes will be discussed.The main results are listed as followed.1.Phenotypic variation analysis within 179 Chinese cabbage genotypes indicated that the variation coefficient of heading types is the highest,with value of 66.4%.There are eight traits with variation coefficient above of 40%,in ordering with transection shape of middle head vein,head performance outside,shape of short stem inside,vertical diameter of short stem inside,heading shape index,color of middle head vein,plant weight,inside leaf color of head.Head weight has also high variation coefficient,with value of 39.2%?However,the variation coefficients of heading characteristics,leaf width,leaf length and plant width were low.2.By association mapping of 33 phenotypic traits and 161 InDel and SSR markers,36markers significantly(P?10-5,R2>0.1)associated with 20 traits were found.In 18 very strong associations,one molecular marker A03-2-1 was significantly associated with the shape of warf stem;two markers A10-3-2 and A10-3-3 was significantly associated with bolting;seven markers A01-S156#01,A05-4-2,A05-8-1,A05-8-3,A06-1-2,A06-S24#01 and A10-2-2 were significantly associated with heading type;one marker A06-S24#01 was significantly associated with number of head leaves;one marker A01-S155#03 was significantly associated with width of head and width of leaf;three markers A03-2-2,A04-8-2 and A06-1-2 were significantly associated with color of petiole and midvein;three markers A04-10-2,A06-1-2 and A10-2-2 were significantly associated with heading index;two marker A03-2-2 and A06-1-2was significantly associated with out leaf color;two marker A04-S60#01 and A06-S24#02 were significantly associated with the germination ability of head short stem;one marker A05-8-3 was significantly associated with Petiole shape in transverse section;one marker A04-8-1was significantly associated with diameter of head;one marker A10-2-3 was significantly associated with head habit.In addition,13 markers were detected associations with more than two traits,two of them A03-2-2 and A06-1-2 were associated with outside leaf color and color of petiole and midvein;one of them A06-S24#01was associated with heading type and number of head leaves.Six traits were found associations with more than two markers,in which the triat of heading type had the most associated markers of seven.3.Using DH88,one high density genetic map including 1603 SNPs distributed across10 linkage groups was constructed,with a total length of 1469.5 cM and average distance0.92 cM between markers.Using F2,one genetic map including 135 markers?99 InDel and 36 SNPs?distributed across 10 linkage groups was constructed,with a total length of1145.7 cM and average distance 8.93 cM between markers.There were 36 common SNP markers between the two maps,in which 899 markers in DH88 and 64 markers in F2 were genes specific.The markers order between genetic map and physical map of DH88 and F2was similar.4.QTL analysis indicated that 95 QTLs controlling 26 traits?10 head development,13 leaf development and 3 others?were detected in the used DH88 and F2 populations,distributing across 10 linkage groups.For detected QTLs,48 of them were related to head development traits,37 were related to leaf development,10 of them were related to leafy number,color and hair at rosette stage.On the other hand,we found six chromosome regions co-controlling at least four leaf development related traits,evolving the top of A03,middle of A03,middle of A04,middle-up of A05,middle of A06,bottom of A08 and bottom of A09.And four main chromosome regions controlling head related traits were identified,evolving the middle of A05,middle of A06,bottom of A08 and bottom of A09.Among 85 QTL regions related to leaf and head development,58 possible co-segerating genes were found.In leaf hair QTL,one candidate gene BrGL1?homologue gene BrGL1 in arabidopsis?related to leaf hair development in Chinese cabbage was found,with distance of 163kb to the closest marker.5.Based on the expression levels and time points of 10 candidate genes,the gene BSA-g4 had the highest level,then there were Bra015396,BSA-g1,BSA-g2 and BSA-g3,genes Bra023570,Bra008613,Bra033844 and Bra005465 had also low expressions,the lowest one is Bra002479.The differences in gene expressions of nine genotypes were mainly at developmental stages of 29-60 days after sowing?seedling,rosette and early heading?,in which the highest level was appeared at seedling or rosette stage and the lowest level was at middle-late heading stage?73 and 88 days after sowing?.Eight genes had high expression at 29 and 36 days after sowing?mainly seedling stage?,two genes had high expression at 36 and 40 days after sowing?mainly rosette stage?.6.Based on expression pattern of 10 candidate genes at three developmental stages?seedling,rosette/early heading and middle/late heading?,four expression patterns were observed.Five genes were high-middle-low patter,with high expression at 29 and 36 days after sowing;two genes were high-high-low pattern,with high expression from 29 to 60days after sowing;two genes were high-low-low pattern,with high expression at 29 and 36days after sowing;one gene were low-middle-low pattern,with high expression from 40 to60 days after sowing.7.Based on regulating head development of 10 candidate genes,relative expression of three genes BSA-g1,BSA-g2 and BSA-g4 increased clearly at rosette and early heading stage,maybe being related to head development,one of them BSA-g1 was possibly related to heading type.Relative expressions of two genes BSA-g3 and Bra023570 were high at seedling stage,indicating that early expression could be related to heading type and time.In overlapping and folding types two genes Bra008613 and Bra005465 had high expression for early heading ones,but low expression in bundling center type,indicating they could regulate leaf bending and heading time.Two genes Bra033844 and Bra015396had different expression pattern between different types,one of them Bra033844 could be related to heading early or late for overlapping and folding types.One gene Bra002479expressed low in all genotypes,indicating it was not the main regulator during head development. |
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