Fine Mapping And Cloning Of LI1 Gene Of Ultra-short Fiber In Cotton

Cotton is one of the most important cash crops in the world that is the main source of natural fiber and mainly used in textile industry.Cotton fiber is the main product of cotton and is a single-celled hairy body developed from the outer epidermal cells of ovule.In the process of cotton fiber development,the number of ovule epidermal fiber cells directly affects the yield of cotton,and the fiber quality is affected by the elongation of fiber cells.Therefore,by locating and cloning the genes related to the growth and development of cotton fibroblasts and predicing and validating the function,it has important theoretical and practical significance to systematically explain the mechanism of cotton fiber growth and development and the molecular mechanism of cotton plant morphogenesis.Cotton ultra-short fiber mutant li1 is an ideal material for studying fiber elongation,primary cell wall and secondary cell wall thickening.Research shows that the li1 mutant is an actin gene which is the raw material of the cytoskeleton and plays an important role in the growth of fibroblasts.In this study,the Upland cotton ultra-short fiber mutant li1 was used as the material to identify the LI1 gene by map cloning,and its function in fiber elongation was preliminarily provided.The LI1 gene was located between the molecular markers W260 and W385 about the physical distance of 890 kb by the method of map cloning.According to the latest research progress,Cloning and analysis of LI1 Gene,the full length sequence of LI1 cDNA is 1134 bp,the gene consists of 4 exons and 3 introns,encoding 377 amino acid sequence sequences,a G mutated into T at 192 bp,which resulted in a hydrophilic glycine mutated into hydrophobic valine in amino acid levels.The analysis of LI1 gene homologous sequence and protein structure showed that LI1 gene was a highly conserved actin gene.The functional analysis of LI1 was preliminarily verified by using VIGS technique,the results showed that the phenotype of LI1 was similar to that of li1 mutants,which was consistent with the expected results,it was preliminarily predicted that the gene might be the target gene.Then we cloned and analyzed the promoter sequence of the promoter sequence from li1 mutant of Upland cotton and Island cotton 3-79 showed that the promoter sequence was the same,but the expression of the gene in the two materials was very different,it is inferred that the difference in gene expression between the two parents is caused by post-transcriptional regulation,which may be regulated by specific siRNA.In order to identify the core promoter region of LI1 gene,seven promoter deletion expression vectors linked to Gus reporter gene were constructed,the promoter fragment of 441 bp was found to have strong transcriptional activity after Gus histochemical staining,the promoter sequence can be used in cotton transgenic breeding instead of general promoter sequence.In addition,over-expression vector,CRISPR/Cas9 vector,yeast two-hybrid bait vector and transient expression vector were constructed,which laid a foundation for further improvement of the experiment.This study is mainly a basic study of the LI1 gene.So far,the analysis of LI1 gene promoter sequence has not been reported,Our main research result is that LI1 promoter has strong activity,through onlinesoftware PlantCARE preliminary analysis and related plant expression vector construction to enrich the types of plant inducible promoters and to provide theoretical basis for plant genetic and breeding improvement.