Screening Of Thermo Sensitive Genic Male Sterile Lines In Arabidopsis Thaliana

In order to meet the growing demand for food,the two-line breeding system is widely used in many crops.The most widely used male sterile lines in two-line breeding system are thermo-sensitive and photoperiod sensitive genic male sterility.However,the safety of seed production in two-line system may be affected by extreme weather caused by El Nino phenomenon.One of the important directions of Hybrid Rice research in the future is to explore and create more thermo and photoperiod related plant resources and simplify breeding procedures.With Arabidopsis thaliana as the research object,the fertility transformation mechanism of P/TGMS lines has been deeply studied,which is helpful to understand the plant male reproductive system and improves the crop breeding system.The creation of mutants by artificial mutagenesis can help researchers to screen and obtain genetic materials which are difficult to obtain in nature.In this study,the seeds of Arabidopsis thaliana were treated by EMS to produce random gene mutation.The self-bred progenies of the mutated lines were screened according to the degree of fertility from a few to completely none and the degree of fertility from completely none to totally restored after low temperature treatment.A total of 68 sterile lines were obtained.The results of SEM showed that the exine pattern of 28-2 and 29-1 was absent,while the exine pattern of 77-1 was relatively complete.Combined with the results of semi thin cross section of anther,the deposit of sporopollenin in 28-2 was abnormal,the tapetal cells in 29-1 were expanded,vacuolated and pre-degraded and the tapetal development of 77-1 was defective.The results showed that the pollen exine pattern of most lines was normal,but the pollen eventually collapsed;5-5 and 1-2 had the defects of pollen exine pattern and could not form normal grid structure.In this study,we further studied 8-93 mutant which was completely male sterile at room temperature but could restore fertility at low temperature.The results of scanning electron microscope(SEM)showed that the mutant 8-93 had no obvious defects of pollen wall pattern.The results of semi-thin cross section showed that the microspores gradually vacuolated and degraded at the stage 9 of anther development,while most of the microspores could normally develop to mature pollen after low temperature treatment.The transmission electron microscope(TEM)images showed pollen grains of 8-93 were collapsed at the stage 9 during anther development.Abnormal large vacuoles appeared in the cytoplasm of 8-93 mutant at stage 12,while the microspores of 8-93 severe shrinkage occurred at stage 14.8-93 mutant mapping results showed that RVMS893 was located on chromosome 5.The results of high-throughput sequencing confirmed the initial localization.The results of genetic complementarity assay demonstrated that the mutation of RVMS893 led to the phenotype of 8-93.The fluorescence images photographed by confocal showed that RVMS893 were located in the nucleus in anther four layers: epidermis,endothecium,middle layer and tapetum at stage 6-8s and also located in microspore nucleus at stages 9-10 during anther development.It is reported that RVMS893 encodes a t RNA methyltransferase.Combined with the above experimental results,it is speculated that RVMS893 may function by modifying the translation level of some key genes in microspore development during anther development.Further analysis of the function of RVMS893 gene will provide new understanding of the relationship between RNA methylation and male reproductive development in plants.