A kind of Brassica napus nac47 transcription factor and its preparation method and application
A technology of Brassica napus and transcription factors, applied in the field of biological genetic engineering, can solve the problem of less rape
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[0029] 1. Cloning and sequence determination of the cDNA sequence of rapeseed NAC47 gene.
[0030] According to the sequence information of the sequenced genome of Brassica napus that has been published, two pairs of gene-specific primers are designed, and the coding region of the gene is amplified using Brassica napus as a template. The primer sequences are:
[0031]BnaNAC47-F:
[0032] 5'TTAGGTCATGATAAGCAAGGATCCAAGATCAAG3'; BnaNAC47-R:
[0033] 5'CGCGTCGACGCCTTGATACTTAAGGTGAG3'.
[0034] The total RNA of rapeseed seedlings was extracted with Plant RNAkit (Omega). After quantification by NanoDrop1000, 2.5 μg of total RNA was taken, and the reverse transcription reaction was carried out with RevertAid RNase H-First Strand cDNA Synthesis Kit (Fermentas) according to the instructions.
[0035] 2. PCR reaction system (50μL)
[0036] 5 PrimeStarbuffer 10μL;
[0037] cDNA template: 1 μL;
[0038] dNTPs (10mM each) 1 μL;
[0039] PrimerF (20 μ M) 1 μ L;
[0040] Primer R (20 ...
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