Transcriptome Analysis Of High-Stalk And Dwarf Castor And Functional Studies Of RcBKI1 And RcGASA9 Genes

Castor bean（Ricinus communis L.）is a very important oil crop,known as"green oil"for its high oil content and reproducibility.At present,most castor varieties used are of high-stalk type,which have poor lodging resistance and are not conducive to mechanized harvesting,seriously hindering the development of castor industry.Selection and utilization of dwarf castor is one of the economic and effective measures to solve the mentioned problems.Due to the incomplete genomic information and the lack of expression profile data,the research about dwarf gene mining and dwarfing mechanism of castor is relatively lagging behind,resulting in the lack of mature theoretical and technical achievements for the research of dwarf castor breeding.In order to reveal the regulation mechanism of castor plant height,RNA sequencing（RNA-Seq）technology was used to compare the changes of gene expression in high-stalk and dwarf castor.In addition,plant hormone-related BKI1and GASA genes play an important role in plant growth and development,but they have not been systematically studied in castor.Combined with transcriptome results,the present study preliminarily analyzed the functions of Rc BKI1 and Rc GASA9 genes.The main results are as follows:The expression patterns of genes in high-stalk and dwarf castor were analyzed at transcriptome level.Dwarf 3358 and high-stalk 2129 castor strains were used as parents for hybridization,and the F₂ population showed strain height character separation.Hi Seq2000,a high-throughput sequencing platform,was used for transcriptional level analysis of high-stalk and dwarf castor,and a total of 6 557 differentially expressed genes（DEGs）were obtained.Enrichment analysis of metabolic pathways showed that these DEGs were mainly involved in plant hormone signal transduction pathway.The function of castor Rc BKI1（30170.t000310）gene was initially identified.BKI1 is an important regulatory protein in brassinolide signaling pathway.The complete open reading frame（ORF）of Rc BKI1 is 999 bp,containing only one exon and encoding 332amino acids.According to RNA-Seq results,the expression level of Rc BKI1 gene in dwarf castor was down-regulated by 4.3 times compared with that in high-stalk castor.Through genetic transformation,a total of three positive Rc BKI1 overexpressed castor were obtained,with a transformation rate of 0.07%;based on CRISPR/Cas9 gene editing technology,one Rc BKI1 knockout plant was obtained to be tested.The Rc BKI1 overexpressed castor showed shortened internodes and dwarf phenotype compared with the control group,proving that Rc BKI1 gene is a negative regulator in the development of castor plant height.The function of castor Rc GASA9（29728.t000025）gene was initially identified.GASA gene family responds to the signals of plant hormones such as gibberellin.The complete ORF of Rc GASA9 gene is 342 bp,including 4 exons and encoding 113 amino acids.According to the results of RNA-Seq,the expression level of Rc GASA9 gene in dwarf castor was down-regulated by 2.6 times compared with that in high-stalk castor.Through genetic transformation,a total of two positive Rc GASA9 overexpressed castors were obtained,with a transformation rate of 0.07%;seven CRISPR/Cas9 Rc GASA9 knockout plants were obtained to be tested.Compared with the wild type castor,the internodes and plant height were increased in Rc GASA9 overexpressied castor,indicating the important regulatory function of Rc GASA9 in the development of castor plant height.