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Overexpression of OsCesA4 and RNAi of Three Lignin Biosynthesis Related Genes in Flax

Author: LiXiang
Tutor: ChenXinBo
School: Hunan Agricultural University
Course: Biochemistry and Molecular Biology
Keywords: Flax Bast fibers Lignin Cellulose Genetic transformation system 4CL
CLC: Q943.2
Type: Master's thesis
Year: 2011
Downloads: 54
Quote: 0
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Abstract


Flax (Linum usitatissimum L.) is one of nature one of the three major fiber crops . Belong to the primary phloem fiber flax fiber has a tensile strength , softness and fineness . Because self -pollination , the relatively small genome advantages , flax crops as a mode of research hemp fiber development . In the textile and paper industry , lignin content , the more increase unglued retting process time , reduce the quality of bast fibers , increasing industrial pollution . This study was constructed the cellulose synthase OsCesA4 gene expression vector and flax lignin biosynthesis genes by RNA interference plant expression vector , and flax genetic transformation system fumble optimization , to raise flax cellulose content by means of genetic engineering for the study and ( ? ) laid the foundation for the quality of the content . The results are as follows : 1. Research through the digestion method 3462bp full-length cDNA , obtained from the full-length cDNA sequences contain OsCesA4 the AK100475 's pFLCI plasmid vector , and blunt-end ligation into pCAMBIA1301M expression vector , constructed plants over express the carrier pOsCesA4 - OE . Confirmed by restriction analysis and sequencing plant over - expression vector was constructed correctly . Amplified by PCR the of flax lignin synthase gene CCoAOMT , 4CL , COMT cis - trans fragment used for RNAi . And finally even into the expression vector pCAMBIA1301M , built RNAi expression vector pCCoAOMT - RNAi , p4CL - RNAi and pCOMT -RNAi connected via intermediate vector pB3D-Linker . Optimized flax Agrobacterium-mediated genetic transformation system . Experimental groping determine the best pre- incubation time 10d, dip time of 30 min , co-cultured 3d, the use screening stages of differentiation hygromycin concentration of 10 mg / L , the rooting stage hygromycin use concentration of 15mg / L . 4 . 4CL gene RNAi flax transgenic plants , and positive transgenic plants detected by DNA level . RT-PCR test results showed that 4CL expression down-regulated in the transgenic plants , indicating that the gene expression was inhibited . The expression vector pOsCesA4 -OE , pCCoAOMT -RNAi pCOMT-RNAi transformed flax has been some resistance adventitious buds , but there have been some problems in the subsequent elongation and rooting process , yet did not get the transplant survival of transgenic plants .

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CLC: > Biological Sciences > Botany > Plant Cell Genetics > Plant Genetic Engineering
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