Application of stress-tolerant relative protein MtMYB5 of plant in cultivating stress-tolerant plant
A stress-tolerant plant and protein technology, applied in the field of genetic engineering, can solve problems such as unsatisfactory effects
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Embodiment 1
[0029] Cloning of embodiment 1, MtMYB5 gene
[0030] 1. Acquisition of MtMYB5 gene
[0031] The gene whose expression was significantly up-regulated was found in chip screening of alfalfa truncated A17 after salt stress. In order to study the function of this gene in abiotic stress, 4-week-old seedlings of alfalfa truncated A17 were poured with 250 mM NaCl aqueous solution for one hour. RNA was extracted, reverse-transcribed to obtain cDNA, and the cDNA of truncated alfalfa A17 was used as a template, and MtMYB5___5' and MtMYB5___3' were used as primers to carry out PCR amplification to obtain a PCR product, which was inserted into the pMD18 T-simple vector (purchased from TaKaRa Bioengineering Company), obtained the recombinant vector, sequenced, and the result was that the PCR product had the nucleotide sequence 1 in the sequence listing, and the gene of the PCR product was named MtMYB5, and the coding region of the gene was sequence 1 to 5 in the sequence listing. 1-837 at...
Embodiment 2
[0042] Example 2, Obtaining and Functional Research of Transforming MtMYB5 Arabidopsis
[0043] 1. Construction of pCAMBIA1302-MtMYB5 constitutive high-efficiency expression vector
[0044] Invitrogen synthesized specific primer pairs containing BglII and Spe I restriction sites respectively:
[0045] 5'-AGATCTTATGGATACTAATTACAAAACCAATA-3' (SEQ ID NO: 5)
[0046] 5'-ACTAGTTCATAAATCATCAGCCAATTGTTGC-3' (SEQ ID NO: 6)
[0047] Using pMD18 T-simple-MtMYB5 as a template, using the DNA molecule shown in Sequence 5 and the DNA molecule shown in Sequence 6 as primers, amplified to obtain a fragment containing the restriction sites of BglII and Spe I, which is the same enzyme The cut vector pCAMBIA1302 (Center for the Application of Molecular Biology to International Agriculture, www.cambia.org) was ligated, and the ligated product was transformed into Escherichia coli to obtain a transformant, and the plasmid of the transformant was extracted and sequenced. The results showed that t...
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