Genetic Analysis And Gene Mapping Of White-leaf Trait In Resynthesized Brassica Napus

White-leaf mutants are important materials to study the mechanism of plant chloroplast development.In this study,green-leaf G7097 and G2127 and white-leaf W7105,which are resynthesized Brassica napus were used as materials.Preliminary genetic analysis and gene mapping of white-leaf trait were performed by means of plant microscopy,QTL-Seq and RNA-Seq technique.The results will lay a theoretical foundation for revealing the mechanism of chloroplast development and photosynthesis in rapeseed.At the same time,it provides a theoretical basis for the application of white-leaf traits as a morphological marker for crop breeding.The main results are as follows:1)The white-leaf trait of W7105 is affected by temperature.Their leaves at the seedling stage are normal green.However,after vernalization by low temperature,the newly grown young leaves turn to be white,and only the leaf margins and pod bills are green.If the white-leaf plants are moved into the greenhouse（25℃）,the albino leaves gradually become green.2)The results of chlorophyll content measurement and transmission electron microscopy showed that there was almost no chlorophyll in the white leaf part of 7105.There was little matrix thylakoid and thylakoid structure in the chloroplasts,and vacuolization was observed in the chloroplast structure.Expectedly,the chlorophyll content and chloroplast structure of the leaf edge were as normal as that of the green leaves.3)Genetic analysis showed that white-leaf is recessive to green-leave trait.It is a quantitative trait controlled by multiple genes.By using QTL-Seq technology,two intervals in ChrC03（1,152,377～1,191,666 and 3,027,120～3,508,054）were supposed to be associated with white-leaf trait.Within the given interval,8 markers were developed.However,no QTL was obtained presently by scanning in BC₁ population with these markers.4)The RNA-Seq analysis of the green and white leaves of W7105 and the green leaves of G7097 were performed.The Common differential genes of GL vs WL and WGL vs WL were regarded as differentially expressed genes（DGEs）.There were 2046 DGEs screened by using B.napus as reference genome,1114 of which were up-regulated and 932 genes down-regulated.There were 2414 DGEs screened by using B.rapa and B.oleracea as a genome,1336 of which were up-regulated,and 1078 genes were down-regulated.The KEGG enrichment analysis showed that in the white leaves,there were a large number of genes down-regulated in pathways such as photosynthesis pathway,photosynthesis-antenna protein pathway,chlorophyll synthesis pathway,and carbon fixation pathway,while in ribosome synthesis pathway,RNA degradation pathway and other pathways,a large number of genes showed up-regulated expression.GO function enrichment analysis revealed significant enrichment in chloroplast thylakoids,basal granules,photosynthetic system,and plastid development entries,consistent with transmission electron microscopy observations.The results indicated that defect in chloroplast development may contribute to the failure of chlorophyll synthesis and accumulation,which ultimately leads to white-leaf phenotype.5)A total of 174 annotated genes existed in the candidate interval on Chr03.In combination with the transcriptome data,three genes were found to be differentially expressed at the transcriptional level.Only one gene Bol034281 with the expression level1.32-fold lower in white-leaf was localized in the chloroplast.The gene is involved in coding a S13 subunit of 30S ribosome,thus it may be a candidate gene for white-leaf phenotype.