| The main component of cotton fiber is cellulose,the fibers of cotton are lint and fuzz.The study on the function of cotton fuzzless gene can explain the mechanism of cotton fiber formation.The multi-omics combined analysis method can effectively discover the functional genes related to fuzz,and analyze the formation mechanism of fuzz from the molecular level.In this study,a hybrid population of upland cotton Xinluzao50 and upland cotton fuzzless mutant Xinluzao50 FLM was constructed,BSA sequencing analysis was used to identify the chromosomes and candidate regions associated with fuzz development,using the electron microscope and paraffin section,we observed the different period of fuzz development in two upland cotton materials,and simultaneously sequenced the transcriptome,and combined with the location interval of BSA analysis,screened out the candidate genes,finally,the function of candidate genes was verified by VIGS technique.The main results were as follows:（1）The field character statistics of F1population showed that the phenotype was lint-fuzzless.Then,from 2019 to 2021,a survey was conducted on the traits of F2population without fuzz,and the number of investigated plants were 320,1167 and 1164,in which the number without fuzz and with catkins accounted for 74.4%,73.4%and 73.6%of the total number of plants,respectively.Through the paraffin section experiment,it was found that the 3 DPA of the fuzz in the two materials was different.（2）By BSA sequencing analysis,3 chromosomes were eventually associated with 6 candidate regions,with a total length of 10.61 Mb,containing 506 genes.In the candidate region,there were 30non-synonymous mutant genes among the parents.（3）A total of 917/620,1105/1803,328/348,257/535 and 457/267 genes were up-or down-regulated by transcriptome sequencing in the Xinluzao 50 FLM and Xinluzao 50,respectively.Three profiles,profile0,profile19 and Profile12,were identified from the two materials by further identification of differential gene expression patterns.The differentially expressed genes in the two materials were analyzed by WGCNA,and two specific modules closely related to the absence of fuzz were obtained.The 20 genes with the highest connectivity in the MEblack module were selected as the hub genes.（4）By comparing the differential genes of the two materials at 3 DPA and combining with 30non-synonymous mutant genes in the interval associated with BSA,4 differentially expressed genes were screened out in the candidate region.（5）The four candidate genes were analyzed by q RT-PCR,and it was found that the expression of GH＿A05G0946 in the two parents had the greatest change compared with other candidate genes during the development of fuzz.Therefore,the function of this gene was verified to determine whether it was related to fuzz.Bioinformatics analysis was performed on the gene family of the candidate gene,which belongs to the DVL family.The silencing system of GH＿A05G0946 gene was constructed.The results showed that the fuzz of the three lines decreased significantly after silencing.In this study,four candidate genes were obtained through the combined analysis of BSA-seq and RNA-seq,and VIGS verification was conducted on candidate gene GH＿A05G0946.The results showed that GH＿A05G0946 may be involved in the formation of upland cotton fuzz. |
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