Gene for regulating polar wood yield and application thereof
A gene, poplar technology, applied in the R2R3-MYB gene PagMYB199 and its application fields, can solve the problems of limited transformation effect, and achieve the effect of great research value and application potential
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Embodiment 1
[0021] Example 1 poplar R2R3-MYB gene PagMYB199 molecular cloning
[0022] 1. Poplar PagMYB199 gene cloning
[0023] Take the poplar stems grown in the culture room for about 6 months, use the CTAB method to extract the total RNA, and use the RT-PCR technology to amplify PagMYB199 The full-length cDNA of the gene was connected into pMD18-T for sequencing (see SEQ ID No.1). The amplification conditions were: pre-denaturation at 95°C for 5min; 35 cycles of 9430s, 40s at 55°C, 1min at 72°C; fragment extension at 72°C for 8min. The amplification primers are:
[0024] PagMYB199cDNAF:5'-ATGGGCAGACAACCTTGTTG-3';
[0025] PagMYB199cDNAR:5'-TTAATGCTTGCCACCCATGTC-3';
[0026] 2. Poplar PagMYB199 Gene sequence information and characteristic analysis
[0027] PagMYB199 The gene contains a coding region of 831bp (see SEQIDNo.1), the molecular weight of the encoded protein is 30.7kDa, and there is a R2R3-MYB conserved domain consisting of 96 amino acids at the N-terminus, which is ...
Embodiment 2
[0030] Example 2 poplar R2R3-MYB gene PagMYB199 transgenic application
[0031] 1. Poplar PagMYB199 Gene overexpression and construction of antisense suppression vector
[0032] poplar PagMYB199 The full-length cDNA of the gene was forwardly and reversely connected to the pCAMBIAL1300 vector, and after the sequence was correct, it contained PagMYB199 Plant overexpression and antisense suppression vectors for genes (see figure 2 ).
[0033] 2. Agrobacterium-mediated transformation of poplar
[0034] Select hybrid poplar leaves that have grown for about 4 weeks and are in good condition, remove the main veins and leaf edges, cut them into leaf disks of the same size, and put them into the differentiation medium for pre-cultivation in the dark for 3 days. At the same time, will PagMYB199 The overexpression and antisense suppression vectors were transformed into Agrobacterium EHA105 and cultured at 28°C until OD 600 =1.0-1.3, prepared as an infection solution. Immerse th...
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