Drought resistant relevant protein IbBT4 and coding gene and application thereof
A technology encoding genes and proteins, applied in drought-resistance-related protein IbBT4 and its encoding genes and applications, can solve problems such as extinction, solute leakage, and threats to agricultural safety
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Embodiment 1
[0076] Embodiment 1, the acquisition of IbBT4 gene
[0077] 1. Acquisition of cDNA template
[0078] The total RNA of the sweet potato line Xushu 55-2 test-tube plantlets was extracted with a plant total RNA extraction kit, and the first-strand cDNA was reverse-transcribed from the total RNA with a PrimeScriptTM RT reagent Kit with gDNA Eraser kit.
[0079] 2. Using the cDNA obtained in step 1 as a template, design and artificially synthesize primers 3GSP-1 and 3GSP-2 according to the EST sequence, use the RACE method to amplify a 3′-RACE fragment of about 800bp, and clone the 3′-RACE fragment and The vector pMD19-T was ligated to obtain recombinant plasmid 2. The recombinant plasmid 2 was sequenced to obtain the nucleotide sequence of the 3'-RACE fragment. The primer sequences are as follows:
[0080] 3GSP-1: 5′-CGAAAACTTTAGGCAGCAGG-3′
[0081] 3GSP-2: 5'-AGTCCGTTTCCTTCACCGAA-3'
[0082] 3. Design and artificially synthesize primers 5GSP-1 and 5GSP-2, use the cDNA obtain...
Embodiment 2
[0089] Example 2. Application of IbBT4 protein in improving plant drought resistance.
[0090] 1. Construction of recombinant plasmid pCAMBIA super1300-IbBT4-GFP
[0091] 1. Artificially synthesize the double-stranded DNA molecule shown in positions 1 to 1116 from the 5' end of SEQ ID NO.2. Using the double-stranded DNA molecule as a template and OE-F-Xba I:
[0092] 5′-GC TCTAGA ATGGGTAAGCTTTCGGATTC-3' (underlined is the recognition sequence of restriction endonuclease Xba I) and OE-R-Pst I: 5'-AA CTGCAG TGTTGCTTCAACTGAGAAAAAT-3' (the underline is the recognition sequence of the restriction endonuclease PstI) is used as a primer for PCR amplification to obtain a double-stranded DNA molecule containing the restriction endonuclease XbaI at the N-terminus and the restriction endonuclease PstI at the C-terminus .
[0093] 2. Digest the vector pCAMBIA super1300-GFP with restriction endonucleases Xba I and Pst I, and recover the vector backbone 1 of about 10783 bp.
[0094] ...
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