Fine Mapping The Rice Curled Leaf Gene CL And Improvement The Molecular Marker That Used For The Chromosome Segment Substitution Lines Derived From "Peiai64S/93-11"

The cl mutant was derived from the anther culture of an autotetraploid rice hybrid. It will be helpful to understand the development program of the rice leaf by cloning the mutant gene.This study was to determine.,the agronomic and genetic characteristics, genetic mechanism of the mutant,to map the gene and to establish the foundation for cloning and functional analysis of the gene.The main results as follows:Before tillering,the cl mutant looked normal,After tillering,phenotypes comparison between cl and wild type indicated that the plant height,the number of primary and secondary branches of cl was significant lower than that of brother lines of wild type.but the tillering numbers are more than the later.The panicle of cl is shorter and it's grain number is smaller,and grain percentage is lower.Genetic analysis show that the phenotype of curled leaf in the cl is controlled by a single recessive nuclear locus,CL.To map the CL locus,we generated F₂ mapping population derived from a cross of the cl mutant and the cultivar ZhenXian97B.We mapped the cl to an interval between markers RM60 and RM22 on Chromosome 3.To narrow down searching for a candidate gene affected in cl,A large F₂ mapping population consisting of more than 8,000 plants,of which 1800 segregants showing the cl mutant phenotype were used for fine-mapping,between markers RM60 and RM22,we further developed two SSR markers and four InDel markers,the cl locus was finally located in a 23-kb DNA region between SSR marker RM523 and InDel marker g62 on a single BAC098695.Within this region,two opening reading frames were predicted,the first ORF encodes a fizzy-related protein.the second ORF encodes an RNA binding protein.To define the molecular lesions of cl mutant,the two ORFs corresponding to the putative CL gene from wild type and cl were amplified by PCR and were sequenced.Comparison of the sequences revealed that only the first ORF carries two nucleotide mutations in codon 38(C deletion) and in codon 40(G→C) in the first exon.which results in.Therefore,we tentatively designated the first ORF as the CL gene.Through RT-PCR,We amplifed the full cDNA of CL,and know the structure of gene, containing ten exons and nine introns.We amplified a 6.2kb full length DNA sequence of CL.The identity of CL was subsequently confirmed by genetic complementation experiments.