| Rice is the principal dietary source of carbohydrates in the world.Amylose is the primary component in grains.Its accumulation in caryopsis filling process is related to expression of Wx gene.In this research,CRISPR/Cas9 was used to edit this gene to generated rice mutant varieties with amylose varying levels.(1)There were totally 16 allele mutants which assorted four types including wxpr being bases substituted on the core promoter region,deleted bases in 5’-UTR of wxu,and wxe2 as well as wxe3 large fragment deficiency in exon 2 and exon 3,respectively.(2)The starch aspect of Wx allele mutants was observed.The AC of wxpr-1lessened to 14.09%.In allele mutants wxu-1,the amylose content went down to 11.76%.Meanwhile,the starch grains structure of wxu-1 were loose round and others had no difference.AC in wxe2-1 and wxe3-2 were reduced to 0-2%.(3)The changes of total starch and amylose during seed filling stage of allelic mutant showed that starch accumulation period of wxpr-1,wxe2-1 and wxe3-2 did not disparity,and the starch accumulation rate of wxu-1 was more gentle than wild-type.There was no rapid accumulation period of amylose.(4)Transcriptional level analysis denoted that genes with starch biosynthesis process.There were down-regulations in Wx of wxpr-1,meanwhile 2 folds upregulations of expression in Flo5.In comparison to wild-type,expression of isoamylase was abnormally elevated in wxu-1.For expression of amylopectin biosynthesis relevant genes in wxe2-1 and wxe3-2 were up-regulated,especially SBE3 and ALK.(5)To survey of grains property,the rigidity of brown rice with wxpr-1 had no distinct imparity,nevertheless,it reduced in wxu-1.Compression resistant of wxe2-1increased slightly and GC boosted apparently.The stiffness of brown rice in wxe3-2 did not change. |
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