| Nitrogen is the most important mineral nutrient in plant growth and development.Rice can directly absorb inorganic and organic nitrogen from soil.The uptake of inorganic nitrogen by rice roots was mediated mainly by nitrate transporters and ammonium transporters,which were then assimilated into amino acids in the plasmids of rice roots.Some of the amino acids synthesized in plants or absorbed by the root system from the soil are immediately metabolized,and some of them are temporarily stored or transported to the growing parts or library organs of plants through xylem and phloem,so as to meet the growth needs of rice.Amino acid transporters(AATs)play an important role in these transmembrane transport processes.Many studies have shown that the key regulatory factors in plant growth and metabolism are amino acid transporters,which play an indispensable role in plant growth and development.Rice is the staple food of more than half of the world’s population.The yield of rice is directly determined by the number of panicles,and the number of panicles depends on the number of tillers.Therefore,improving the number of tillers of japonica rice is an urgent need for breeding and producing high-yield and high-quality japonica rice varieties.Preliminary results show that aromatic and neutral transporter(ANT)and Amino acid transporter like(ATLa)were responsive to tiller buds of rice and played an important role in the growth and development of rice.In Arabidopsis Thaliana,only AtANT1 gene of ANT family has been reported.AtANT1 can transport not only aromatic amino acids,neutral amino acids and arginine,but also indoleacetic acid and 2,4-dichlorophenoxyacetic acid.In rice studies,reports suggest,The expression level of OsATL5 in rice was higher.The expression of OsANT3 and OsANT4 was up-regulated when rice was subjected to abiotic stress,but the mechanism of these genes in plant growth and development has not been reported.In this paper,the bioinformatics analysis of OsANT1,OsANT2,OsANT3℃ OsANT4,OsATL5 and OsATL7 was carried out,and Their different response modes to low and high ammonium were analyzed.The mutants of OsANT1,OsANT2,OsATL5 and OsATL7 genes were analyzed in detail.Gene knockout materials were constructed by using CRISPR_Cas9 technology.The differences of tiller,biomass,panicle shape and yield between wild type and mutant were analyzed and compared.The main findings are as follows:1.Phylogenetic tree was constructed based on the protein sequences of each gene to analyze the phylogenetic relationship between rice and Arabidopsis Thaliana.In rice,the structure of ANT1 and ANT2 genes was similar,and the protein sequence consistency reached 52.58%,which was highly related to ANT1 of Arabidopsis Thaliana.The structure of ANT3 and ANT4 genes in rice is similar,the protein sequence consistency of ANT3 and ANT4 was the highest in rice,up to 80.28%,which was highly related to ANT2,ANT3 and ANT4 of Arabidopsis Thaliana.The structure of ATL5 and ATL7 genes in rice is similar,The protein sequence consistency of ATL5 and ATL7 in rice is 36.85%,ATL5 of rice and ATL2 and ATL3 of Arabidopsis Thaliana have high affinity.ATL7 of rice has high affinity with ATL4 and ATL5 of Arabidopsis Thaliana.The protein domain analysis of each gene in rice showed that they all had typical transmembrane structures and predicted their function on the plasma membrane.2.qRT-PCR was used to detect the different responses of different genes at the rhizome junction to high and low ammonium.OsANT1 was inhibited expression by high and low ammonium;OsANT2 was induced expression by high ammonium;OsANT3 was induced expression by low ammonium;OsANT4 was inhibited expression by low and high ammonium;OsATL5 was inhibited expression by high ammonium;OsATL7 was induced expression by high ammonium.3.Field data showed that the knockout of OsANT1 had no significant effect on panicle shape of rice,but could significantly reduce the number of tillers by about 16%.Significantly reduced rice yield by about 39.9%.4.Field and hydroponic data showed that the knockout of OsATL5 had no significant effect on tiller and biomass of rice.5.Field data showed that the knockout of OsATL7 had no significant effect on the panicle shape of rice,but could reduce the number of tillers by about 25.9%and significantly reduce the yield per plant by about 36.4%. |
