Cloning Of Ct Region Of Acetyl-coenzyme A Carboxylase Gene From Avena Sativa L And Establishment Of CRISPR/Cas9 Technology System

Avena sativa L belong to Avena L,an annual herb that is an important food and forage,feed crop.The emergence and widespread use of chemical herbicides have resulted in effective and labor-saving methods for field weeding.“Na bu”,also known as sethoxydim,is a selective herbicide.When “Na bu” is used to remove the grass weeds in the Avena sativa L field,it will cause harm to the Avena sativa L and limit the removal of the weeds from the herbicides.Acetyl coenzyme A carboxylase(ACCase)is a key enzyme that catalyzes the synthesis of fatty acids in plants during plant metabolism.It is a target protein for the action of herbicides.It recognizes the protein structure of ACCase and is associated with it.In combination,the purpose of inhibiting ACCase activity is achieved,so that the lipid metabolism is hindered,and then the grass weeds are killed.This study cloned the CT region sequence of ACCase of sweety 1 and performed bioinformatics analysis.CRISPR/Cas9 gene editing vector was constructed and the vector was introduced into mature sweety 1 embryo via particle bombardment method.In order to obtain anti-"Na bu" new Avena sativa L germplasm.The study yielded the following results:1.Using the method of homologous sequence cloning,the 2188 bp region of the CT region of Avena sativa L ACCase gene was obtained,and wheat(Triticum aestivum),Alopecurus japonicus,Alopecurus myosuroides,etc.have been reported.The ACCase sequence homology is 100%.And the 1769 th amino acid sequence in the sequence is leucine,which is sensitive to net acquisition.2.Using CRISPR/Cas9 technology to construct an anti-acquired net expression vector.3.53 strains of plants transformed by particle gun were obtained.After positive identification and analysis,two mutants of Cas9 were generated.All the deletions were small fragments.The deletions were random and indirect deletions.However,the correct integration of the repair template was not detected in the two mutant transgenic plants.4.The conversion efficiency using the gene gun method in this experiment was 3.8%.For the first time,this study used CRISPR/Cas9 gene editing technology to edit the oat genome,providing experimental basis for the application of gene editing technology in oat molecular breeding.