Adverse resistance related protein IbBBX24 and coding gene and application thereof
An ibbbx24, coding technology, applied in the fields of application, genetic engineering, plant genetic improvement, etc., can solve inter-species and intra-species hybrid incompatibility, limit the utilization of sweet potato breeding resources and the free combination of parents, and it is difficult to breed high-quality, New varieties of sweet potato with high yield and resistance to vine-cutting disease, etc.
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Embodiment 1
[0085] Embodiment 1, the acquisition of IbBBX24 gene
[0086] The steps of obtaining the IbBBX24 gene are as follows:
[0087] 1. Obtaining the template
[0088] The total RNA of the young leaves of the sweet potato salt-tolerant mutant ND98 was extracted with a plant total RNA extraction kit, and the total RNA was extracted with PrimeScript TM 1st Strand cDNA Synthesis Kit reverse transcribes the first strand cDNA.
[0089] 2. Construct the cDNA-AFLP subtraction library, and obtain the EST sequence shown in sequence 4 in the sequence listing. According to the nucleotide sequence of EST sequence, primers GSP-1 and GSP-2 were designed and synthesized artificially.
[0090] 3. After completing step 2, use the cDNA obtained in step 1 as a template, use the GSP-1 and GSP-2 synthesized in step 3 as primers, and use the RACE method to amplify a 3'-RACE fragment of about 700 bp, and convert the 3'- The RACE fragment was connected with the cloning vector pMD19-T to obtain recombi...
Embodiment 2
[0099] Embodiment 2, the application of IbBBX24 protein in regulating the stress resistance of sweet potato
[0100] 1. Construction of recombinant plasmids
[0101] A, construction of recombinant plasmid pCB-IbBBX24
[0102] 1. Artificially synthesize the double-stranded DNA molecule shown in Sequence 1 of the Sequence Listing. Using the double-stranded DNA molecule as a template, OE-F-XbaI: 5′-GC TCTAGA ATGAAGATTCAGTGTGATGTGTGT-3' (underlined is the recognition sequence of restriction endonuclease XbaI) and OE-R-SacI: 5'-C GAGCTC TCAACCGAGATCAGGGACAGT-3' (the underline is the recognition sequence of restriction endonuclease SacI) is used as a primer for PCR amplification to obtain a double-stranded DNA molecule containing restriction endonuclease XbaI at the N-terminus and restriction endonuclease SacI at the C-terminus.
[0103] 2. Ligate the double-stranded DNA molecule containing the restriction endonuclease XbaI at the N-terminal and the restriction endonuclease Sac...
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