Cloning And Function Analysis Of Glabrous Leaf Gene GLR3?GL5 And Fine Mapping Of Hairy Leaf Gene HL6 In Rice

Rice is one of the important food crops in China,as well as a model plant for functional genomics research in crops.Therefore,it is of great significance to isolate the important morphological and agronomic genes and explore its evolution pathway.Rice trichome is an important agronomic and morphological character,and little information has been known about the rice trichome gene and involved molecular mechanisms.In this study,genes conferring trichome formation were cloned or fine mapped using unique rice genetic germplasms,and also molecular mechanism analysis were conducted.The main results are as follows:Firstly,the glabrous hull gene GL5(Glabrous Leaf 5)were fine mapped to a 33 kb region,in which there were four predicted candidate genes.Genome sequence and qRT-PCR analysis showed that LOC_Os05g02730 was the most possible candidate gene.In addition,the trichome of RNA interference transgenic plants entirely disappeared.These indicated that LOC_Os05g02730 was the causal gene of GL5 and the lack of trichome was caused by the knock down of GL5 expression level.Secondly,based on the allelism tests between GL5 containing material HGL and other 69 glabrous hull materials,we isolated a novel gene controlling glabrous leaf gene GLR3 {Glabrous Rice 3)from WD17993 and fine map it to a 21 kb region and only one gene was predicted.Transgenic analysis showed that RNAi lines had reduced or disappeared trichome on the leaf blades and grain hulls compared with control plants.GLR3 transgenic complementation and overexpression lines showed appeared trichome,but trichome number and length were less than the wild type plants(Nipponbare)with unkown reseaons.GLR3 encodes a SPL domain containing transcription factorlocated in nucleus.qRT-PCR and GUS staining showed that GLR3 widely expressed in different tissues,and with less expression level in the stem of Nipponare and WD17993.Transcriptional activation activity assay showed that GLR3 containing a transcription activation domain from 256 to 427 amino acids.In order to explore the molecular mechanism and interaction between GL5 and GLR3,a series of experiments were conducted.We found that the expression level of GL5 significantly increased in GLR3 RNAi lines but reduced in GLR3 complementation and overexpression lines.Moreover,transient promoter activity assay conducted in tobacco showed that GLR3 repressed the promoter activity of GL5.These results imply that GLR3 may play as a transcriptional repressor of GL5 in the development of trichome of rice.Lastly,a gene controlling villous formation in leaf HL6(Hairy Leaf 6)was fine mapped in a 39 kb region using 19635 BC3F3 individuals derived from the cross between JOAH(Hairy leaf)and Nipponbare.According to the annotation from MSU,there exited four candidate genes.Compared with the Nipponbare,LOC_Os06g44790,LOC_Os06g44810 and LoC_Os06g44820 had nucleotide polymorphisms in JOAH,and LOC_Os06g44790 showed higher expression level in JOAH.But the complementation and RNAi transgenic plants of these three genes showed no phenotypic change.The exploration of the unkown reason is ongoing.