Экспрессионные векторы для редактирования генов кафиринов (k1C5, gKAF1) и гена ameiotic 1 у сорго с помощью агробактериальной трансформации

Abstract

One of the effective approach to improve the nutritional value of sorghum grains is the production of mutants with partially or completely blocked synthesis or the altered amino acid composition of spare proteins - kafirins. The usage of the genomic editing method allows to solve this problem by introducing mutations into the nucleotide sequences of the genes α- and γ-kafirins (k1C5 and gKAF1). Another important area in the genetics and selection of agricultural plants is the development and use of apomixis technologies to fix heterosis in F1 hybrids. The DYAD arabidopsis gene and its homolog ameiotic 1 are the potential apomixis candidate genes involved in the normal course of meiosis, the induction mutations in which is one of the way to solve this problem. In the course of research, the genomic targets (20 bp sequences) were selected to introduce mutations into the genes of α- and γ-kafirins of sorghum and ameiotic 1 gene through CRISPR/Cas genomic editing technology. Guide RNA design is performed using bioinformatic programs CRISPROR and CHOPCHOP. Cloning of the α- and γ- kafirins and ameiotic 1 targets of sorghum into vector pBAtC was carried out at sites AarI. Validation of the success of target cloning was performed by DNA sequencing. The created constructions were transferred by electroporation to the agrobacterial strain AGL0. Currently, the created vectors are used for the agrobacterial transformation of sorghum plants.