Regulation Of GhOKRA And GhPIN3 Genes To Create New Leaf Shape Germplasm Of Gossypium Hirsutum L.

Cotton is not only an important natural fiber crop,but also an important source of vegetable oil and protein.Cotton has become one of the most important cash crops in the world because of its high economic value.Leaf shape is one of the important agronomic traits of cotton.Different leaf shape affects photosynthesis and small field environment,and then affects cotton yield,flowering time,insect resistance and so on.At present,the broad leaf types of cotton varieties used in Chinese production are mainly broad leaf types.Broad leaves have greater potential for single plant production because of their greater leaf area coefficient.Compared with normal leaves,okra leaves have better ventilation and light transmittance,and have a certain degree of resistance to pests in small field environment.However,because of the smaller leaf area coefficient,the yield of okra cotton is generally lower than that of broadleaf cotton.At present,Xinjiang is the main cotton producing area in China,and the planting density is 4-5 times that of inland cotton area.It is of great significance to select and cultivate cotton varieties with proper leaf shape.Previous studies have shown that the GhOKRA gene controls the absence depth of cotton leaves,and the mutation of this gene causes cotton leaves to change from okra leaf type to broadleaf type.In this study,the GhOKRA gene knockout and interference vector was constructed to obtain cotton materials with different leaf shapes,so as to provide materials and theoretical basis for shaping cotton ideal plant type.There are many genes involved in the regulation of plant type.As an auxin transporter,PIN gene family is involved in the development of leaf branch and the formation of leaf shape.In this study,GhPIN3 knockout vector was also constructed to explore its effect on leaf shape and leaf branch.The main results are as follows:（1）The results of measuring the expression of GhOKRA gene in different tissues and at different stages of the leaves of super-okra showed that the D genes of GhOKRA were mainly expressed in shoot tips and leaves,while the A genes were not expressed.At the fourth true leaf stage,the expression of D genes began to increase,and the leaf shape starts to change.（2）The GhOKRA gene editing knockout vector was constructed,and the GhOKRA mutants were created by transgenic technology.A total of 5 GhOKRA transgenic positive strains were obtained,and 5 transformation events were performed,and the leaf shape of the leaves was changed from super-okra leaf to broad leaf.（3）According to the editing effect and type statistics of GhOKRA sub-okra transgenic plants,it was found that there was 1bp deletion in sgRNA1,while there was no editing in sgRNA2.（4）By constructing GhOKRA RNAi interference vector and using transgenic technology to interfere with GhOKRA gene at RNA level,34 positive strains and 21transformation events were obtained,and the leaf defect degree of the transforts was weakened to different degrees compared with that of the super-okra.（5）The knockout vector of leaf branch and leaf type related gene GhPIN3 was constructed,and GhPIN3 mutants were created by transgenic technology.A total of 18transgenic positive plants were obtained,which belonged to 11 transformation events.The leaf edges of these 18 regenerated plants were curled and shrunken upward and inward,the plant type was compact,and the number of leaf branches was reduced.The leaf of T₁generation plants were curled upward and inward,and the Angle between fruit branches decreased,the number of leaf branches decreased,and the plant type was more compact.GhPIN3 not only affects the formation of leaf branches,but also affects the leaf shape.