Effects Of Lipid Transfer Protein OsLTPL18 On The Growth And Development Of Grains In Rice

Nonspecific lipid transfer proteins(ns-LTPs)are widely distributed in the plant kingdom.They can affect plant growth and development and stress adaptation.There are 52 ns-LTPS family members in rice,among which the role of OsLTPL18 in rice growth and development has not been reported.Based on CRISPR technology,knockout mutants of OsLTPL18 was created.It was found that OsLTPL18 knockout mutation significantly affected the grain thickness and grain weight of mature rice.It lays a foundation for further study on the role and mechanism of OsLTPL18 in rice growth and development.The main results are as follows:1.Characteristics of OsLTPL18 gene and its coding protein in rice were analyzedOsLTPL18 is located on chromosome 1 of rice,with a total length of 1785 bp and two exons.The total length of CDS sequence is 384 bp.OsLTPL18 protein contains 173 amino acids with an isoelectric point Pi of 4.36.Its N-terminal has a signal peptide containing 28 amino acids and 8 conserved cysteine(Cys)residues.2.Expression patterns of OsLTPL18 in different tissues and organs of rice were clarifiedThe results of fluorescence quantitative PCR showed that OsLTPL18 was expressed most in young panicles at P6 stage,more in young panicles and stems of seedlings at P5 stage,less in young panicles at P4 stage,grains and roots of seedlings on the third day after flowering and fertilization,and no expression was detected in other tissues and organs.3.Knockout mutants of rice OsLTPL18 were createdBased on CRISPR technology,three types of OsLTPL18 knockout mutants were successfully created.The mutation types were-C,-CT and-T respectively.These three types of amino acid mutations all destroyed the protein domain of OsLTPL18.4.OsLTPL18 significantly affected grain development of riceThe results of trait measurements at maturity showed that OsLTPL18 knockout mutation significantly affected the plant height,panicle length,seed setting rate and grain weight of rice.Among them,compared with the wild type,the OsLTPL18 knockout mutation significantly reduced 1000-grain weight of rice,and the decrease range of 1000-grain weight of three mutants was 10.5%-13.9%.Compared with the wild type,the grain length of OsLTPL18 knockout mutant was not significantly different,but the grain width and grain thickness were significantly reduced,which may be an important reason for the significant reduction of 1000-grain weight of OsLTPL18 knockout mutants;Among them,the decreasing range of grain width is4.57%-5.85%;The decreasing range of grain thickness is 9.07%-9.78%.5.OsLTPL18 knockout mutation significantly reduced seed vigor,germination rate and seedling growth of riceCompared with the wild type,the seed vigor of OsLTPL18 knockout mutant was significantly reduced by 22.53%-33.80%,the germination rate of mutant seed was significantly reduced by 30%-54% at the 72 nd hour after germination,and the plant height of mutant seedling grown at the 7th day was significantly reduced by34.26%-43.86%.6.OsLTPL18 knockout mutation may also affect rice qualityWe found that OsLTPL18 knockout mutation caused the aleurone layer of mutant rice grain to become significantly thinner,the total starch content of grain decreased significantly,with a decrease range of 1.44%-2.23%,and the expression of some starch synthesis related genes and storage protein coding genes in grain changed significantly.It shows that OsLTPL18 knockout mutation is likely to affect rice quality,which needs to be further explored.