Study On Downstream Gene Expression And Lignin Content Of PpMYB4 Transgenic Rice

Pennisetum purpureum is the most common perennial herb in South China.However,its high lignin content in vivo affects its development and utilization.The PpMYB4 is a transcription factor that can inhibit the production of lignin in the body.The cloned sequences had been submitted to NCBI,with the nucleotide accession number of KU612217.In this study,the PpMYB4 transgenic rice was obtained and used for the gene function analysis.The effect of lignin was analyzed to explore the function of this transcription factor.The lignin content and its downstream gene expression in different stages were observed and measured in transgenic rice plants of the PpMYB4,and the effects of the transcription factor on rice lignin were explored.The results are as follows:(1)Overexpression vector of pCUbi1390-PpMYB4 was constructed and transferred into rice by Agrobacterium-mediated method.Transgenic plants were identified by PCR and RT-PCR.Compared with the wild type,the transgenic rice showed a slow growth rate,slender stems,poor support,easy lodging,and yellow-curled leaf tip.(2)Phloroglucinol staining was carried out in the penultimate or antepenultimate internode of the transgenic and wild type rice at heading,flowering and milk ripening stages.The results showed that the staining degree of the fibrous tissue layer under the stem epidermis in transgenic rice was lighter than that of the wild type rice at the heading,anthesis and milk ripening stages.(3)The determination of lignin content by bromoacetyl method indicated that the lignin contents in stems of transgenic rice had the significant decrease of 16.77%,17.74%and 9.45% at heading,flowering,and milk ripening stages compared with wild-typeplants.This results suggesting that PpMYB4 participates in and inhibits the lignin synthesis of transgenic rice.(4)The expressions of key enzyme genes related to lignin synthesis in transgenic rice were detected by qPCR.The relative expression of PAL,C4 H,CAD,CCoAOMT,4CL,COMT,CCR and HCT in the flowering and milk mature transgenic rice was lower than that of the wild type,and the expression of PAL was significantly lower than that of the wild type.The results showed that overexpressing PpMYB4 could regulate lignin biosynthesis by regulating the expression of key genes in transgenic rice.