Targeted Editing Of Maize Photoperiod Sensitive Genes ZmCCT10 And ZmCCT9 By CRISPR/Cas9 Technology

Growth period is an important factor affecting the yield increase and stable yield of maize,and also a hot direction of maize breeding and improvement work.ZmCCT10 and ZmCCT9 belong to CCT family genes,and participate in the regulation of maize photoperiod.ZmCCT10 can inhibit the flowering of maize in long sunshine,knocking out this gene can advance the flowering stage of the maize and improve the multiple cropping index and increase the total annual grain output;knocking out the gene can break the restriction of planting area and expand the planting range and introduce it to high latitude,high altitude and other areas;knocking out the gene can effectively reduce the probability of encountering natural disasters during the growth period and effectively avoid yield reduction.Changing the flowering period of maize and shortening the growth period appropriately are of great significance to ensure the safety and high yield of maize.The results of this study are as follows:(1)ZmCCT10/ZmCCT9-related mutants were successfully obtained by constructing CRISPR/Cas9 knockout vector with single sg RNA.There were four mutations in ZmCCT10 gene: +1T,+1A,C18 T,-2bp,and one mutation in ZmCCT9 gene: +1 A.(2)The flowering period of ZmCCT10-1IT strain and ZmCCT10-1IA strain with KN5585 as the background was obviously 2-5 days earlier in Hainan short-day area and Beijing long-day area,and the plant height,emasculated plant height and spike height were dwarfed by about 20 cm.(3)The ZmCCT9-1IA strain with KN5585 as the background has not yet obtained negative homozygous materials.Preliminary observation on the phenotype of positive materials shows that the plant height,emasculated plant height and ear height are obviously shorter,the mutant ear is obviously smaller than the wild ear,and the overall biomass of the plant is obviously reduced.(4)Through DTM strategy,the late-maturing materials with CML312 as the background obtained two mutant strains related to ZmCCT10 without transgenic elements:ZmCCT10-1IT strain and ZmCCT10-C18 T strain.ZmCCT10-1IT strain,as a frameshift mutant,has a significantly earlier flowering period than the wild type.ZmCCT10-C18 T strain,as synonymous mutant material,has no obvious change in flowering period compared with wild type.