Plant stress tolerance related protein TaSnRK2.10 as well as coding gene and application thereof
A technology for encoding genes and proteins, applied in the field of plant stress tolerance-related protein TaSnRK2.10 and its encoding genes and applications
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Embodiment 1
[0047] Example 1. Isolation of the gene TaSnRK2.10 encoding the stress-resistance-related protein in wheat
[0048] 1. Isolation of TaSnRK2.10, the gene encoding anti-stress-related protein
[0049] 1. Construct the full-length cDNA library of wheat, according to the literature (Mao Xinguo et al., 2005, use the improved Cap-trapper method to construct the full-length cDNA library of I. spelter. Acta Genetics, 32 (8): 811-817) The method described in:
[0050] (1) Total RNA extraction and mRNA purification, wheat total RNA was extracted with TRIZOL, and mRNA was separated and purified with oligo(dT) cellulose.
[0051] (2) Synthesis of first-strand cDNA: Mix 10ug mRNA with primer I, add reagents for first-strand cDNA synthesis after denaturation, when the temperature rises to 40°C, add reverse transcriptase, and when the reaction reaches 40 minutes Primer II was added (first strand synthesis primers below). In order to obtain more full-length cDNA, trehalose and behenitol we...
Embodiment 2
[0075]Example 2, gene TaSnRK2.10 and its application in Arabidopsis
[0076] 1. Acquisition of TaSnRK2.10
[0077] Design primers: upstream primer F1: 5'-CCCAAGCTTGTGGGGGAAGGAAAGGGG-3' (Hind III site, sequence 3), downstream primer RI: 5'-ACGC GTCGACCATAGCATACACTATCTCCCCGCT-3' (Sal I site, sequence 4), wherein downstream primer 3 ' end is located upstream of the gene stop codon.
[0078] The mRNA of common wheat (Triticum aestivum L. Hanxuan No. 10, preserved by the National Crop Germplasm Bank) was extracted and obtained by RT-PCR using 5′-GTGGGGGAAGGAAAGGGG-3′ and 5′-AGTAACATTTGTGGGCGGGC-3′ as primers The full-length cDNA of gene TaSnRK2.10 in wheat;
[0079] Using the full-length cDNA sequence of the gene TaSnRK2.10 as a template, using the above-mentioned primers F1 and R1, the high-fidelity enzyme Pfu was used to amplify the target gene, and a PCR product of 1119bp was obtained, which was sent for sequencing. The result was that the PCR product had the sequence in the s...
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