Study On The Lignin Content Of The Maize (Zea Mays L.) Stalks And Lignin Structure By Downregulation Of ZmCoA

The major constituent of the Maize (Zea Mays L.) stalks are lignin and cellulose. Lignin is a major cell wall polymer of vascular plants that provides mechanical strength and hydrophobicity to vascular vessels. Because of the complex structure of lignin, it is difficult to be effectively used in production. There is negative impact on forage quality, paper manufacturing, and more recently, cellulosic biofuel production. Thus there is significantly economical and environmental value on regulation of lignin biosynthesis by genetic engineering at the molecular level to reduce lignin content or to change lignin components. In this research, maize B73was used as material, using RNAi technology, inhibited the key enzymes in lignin biosynthesis gene expression of ZmCOMTand ZmCoA. We constructed the expression vectors containing cDNAs of COMT and COA transferred into maize mediated by Agrobactirium Tumefaciens. PCR analysis that genes integrated into the genome of the transgenic maize, and analysis the content lignin and cellulose, semi-quantitative RT-PCR analysis, histological observation. The main results are as follows:1、The genomic DNA of maize B73was extracted following CTAB method, the part of the gene COMT and CoA had been amplified by PCR amplification. Using genetic engineering, vector p1301+2F+Bar containing CaMV promoter35S by utilizing pUCCRNAi and p1301vector was constructed. The interference vectors were transformed into Agrobacterium Tumefaciens Ag10by protocol of direct transformation.2、The expression vector p1301+CoA (+)+Bar was introduced into maize zheng58by shoot tip of maize transformation system. And transgenic plants were screened using herbicide.3、T1transgenic plants that tested positive by PCR detection were analyzed according to RT-PCR. The results implied that RNAi reduced the level of ZmCoA expression. Klason lignin and cellulose content of stalks decreased by22.44%, cellulose content increased by23.33%compared with the control.4、Cryostat sections of transgenic lines were observed by microscope in100and200bars. We saw that the color of vascular sheath of transgenic lines stem lighter than the wild-type. This indicated that the ZmCoA gene silencing reduced lignin accumulation in the vascular bundle thus reduced the content of lignin.5、The relative content of lignin monomer by GC-MS was studied. It was found that S/G was0.999in transgenic lines and0.636in wild-type. The G/S increased by57.08%. This indicated that the ZmCoA gene silencing increased S/G, thus facilitated the removal of lignin.In summary, we have successfully builded RNAi expression vectors pl301+CoA (+)+Bar and p1301+COMT (+)+Bar, and conducted genetic transformation by p1301+CoA (+)+Bar. We obtained transgenic lines and made a series of detections, the results were that lignin content in transgenic maize reduced, and structure of lignin changed. This research will provide foundation for breeding of low-content maize and novel insight about lignin.