Plant drought-resistant relevant protein PvSnRK 2.3 and encoding gene and application thereof
A technology related to proteins and plants, applied in the field of genetic engineering, can solve problems affecting wheat yield and quality, and restricting wheat food security, etc.
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Embodiment 1
[0070] Example 1: cDNA Cloning of PvSnRK2.3 Gene Related to Drought Resistance in Switchgrass.
[0071] The 30-day-old switchgrass seedlings were subjected to drought treatment for 5 hours, and the total RNA of switchgrass was extracted with Trizol. 5'RACE kit (5'RACE System for Rapid Amplification of cDNA Ends Kit) (GIBCOBRL, CAT.NO.18374-058) and 3'RACE kit (3'RACE System for Rapid Amplification of cDNA Ends Kit) ( GIBCOBRL, CAT.NO.18373-019) obtained the full-length sequence of PvSnRK2.3 gene 1002bp.
[0072] The total RNA of switchgrass seedlings was extracted with Trizol, and cDNA was obtained by reverse transcription with superscript II (invitrogen) reverse transcriptase. Primers P1 and P2 were designed according to the sequence of the coding region of PvSnRK2.3 gene. Using the cDNA obtained by reverse transcription as a template, PCR amplification was performed with primers P1 and P2. The sequences of primers P1 and P2 are as follows:
[0073] P1: 5'-ATCGTCCAGGAGGCC...
Embodiment 2
[0077] Example 2: Enhancement of plant drought tolerance with PvSnRK2.3 gene
[0078] 1. Construction of recombinant expression vector
[0079] 1) Construction of 35S-PvSnRK2.3 recombinant expression vector
[0080] Using the cDNA obtained by reverse transcription of switchgrass total RNA as a template, PCR amplification was performed with specific primers containing SmaI and SpeI linker sequences; then SmaI and SpeI double-enzyme-digested PCR products were recovered, and the enzyme-digested products were inserted forward into the vector pBI121 Between the SmaI and SpeI restriction sites behind the CaMV35S promoter, the recombinant vector p35S-PvSnRK2.3 was obtained.
[0081] The primer sequences are as follows:
[0082] PvSnRK2.3[SmaI]5'-TC CCCCGG GATGGAGGAGAGGTAC-3'
[0083] PvSnRK2.3[SpeI]5'-GG ACTAGT TCAGTAGGCGTCATCGGCG-3'
[0084] 2. Acquisition and functional identification of transgenic Arabidopsis
[0085] 1) Obtaining transgenic Arabidopsis
[0086] The reco...
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