Sweet potato leaf development and flavonoid enhancement related protein IbBBX29 as well as coding gene and application thereof
A technology that encodes genes and flavonoids, applied in the fields of application, genetic engineering, plant genetic improvement, etc., can solve the problems of lack of genetic resources, incompatibility of intraspecific hybridization, narrow genetic basis, etc.
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Embodiment 1
[0103] Example 1, the acquisition of IbBBX29 gene
[0104] The steps to obtain the IbBBX29 gene are as follows:
[0105] 1. According to the transcriptome analysis, find a highly expressed gene IbBBX29 in sweet potato leaves, obtain the diploid CDS sequence of the gene by comparing with the sweet potato diploid genome database, and synthesize the primer IbBBX29-F:5'- ATGGCGAAAGAAACGAAGAG-3' and IbBBX29-R: 5'-TCAATCAAGCAGTGACGACG-3'. Total RNA from young leaf organs of flavonoid-rich Sushu 33 was extracted with a plant total RNA extraction kit, and the total RNA was extracted with PrimeScript TM 1stStrand cDNA Synthesis Kit reverse-transcribes the first-strand cDNA.
[0106] 2. Using the cDNA obtained in step 1 as a template, using IbBBX29-F and IbBBX29-R as primers to carry out PCR amplification, and ligating the amplified product with the cloning vector pMD19-T to obtain the recombinant vector pMD19-T-IbBBX29. The recombinant vector pMD19-T-IbBBX29 was sequenced to obtain...
Embodiment 2
[0108] Example 2. Application of IbBBX29 protein and its encoding gene in regulating leaf development and / or flavonoid content in leaves
[0109] 1. Construction of recombinant plasmids
[0110] A. Construction of recombinant plasmid pCambia1300-IbBBX29-GFP
[0111] 1. The double-stranded DNA molecule shown in SEQ ID No. 2 of the artificially synthesized sequence listing. Using this double-stranded DNA molecule as a template, IbBBX29-OE-F: 5'-GG GGTACC ATGGCGAAAGAAACGAAGAG-3' (KpnI restriction site is underlined), and IbBBX29-OE-R: 5'-ACGC GTCGAC ATCAAGCAGTGACGACG' (the underlined part is the Sal I restriction site) was used as a primer for PCR amplification to obtain a double-stranded DNA molecule containing the restriction endonuclease KpnI at the N-terminal and the restriction endonuclease SalI at the C-terminal.
[0112] 2. The vector pCambia1300-GFP was double-digested with restriction enzymes KpnI and SalI to recover the large fragment, while the double-stranded DNA...
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