Application of zmbzip68 protein and its encoding gene in regulating low temperature stress in maize
A technology encoding genes and proteins, applied in application, genetic engineering, plant genetic improvement, etc., can solve problems such as limitation, unstable application, limited, etc., to achieve improved tolerance, improved cold resistance, high stability and application effect of value
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Embodiment 1
[0052] Example 1 Construction of homozygous knockout mutant of ZmbZIP68 gene using CRISPR / Cas9 technology
[0053] The ZmbZIP68 gene involved in this example is derived from the maize LH244 inbred line, and its sequence in the maize genome is shown in SEQ ID NO. And positions 4659-6509 are all intron sequences; the cDNA sequence is shown in SEQ ID NO.2, and the amino acid sequence of the encoded protein is shown in SEQ ID NO.1. In this example, the ZmbZIP68 gene was knocked out in maize using the CRISPR / Cas9 technology, and a homozygous knockout mutant of the ZmbZIP68 gene was constructed.
[0054] 1. Construction of recombinant vector pBUE411-pCBC-MTaTb
[0055] After screening, it was determined that CRISPR / Cas9 knocked out two target sequences Ta and Tb of the ZmbZIP68 gene, wherein the target sequence Ta was the reverse complementary sequence of positions 1014-1033 of the sequence shown in SEQ ID NO. Positions 1170-1189 of the sequence shown in SEQ ID NO.2. Design ampli...
Embodiment 2
[0082] Example 2 Low temperature treatment test of ZmbZIP68 gene knockout line
[0083] 1. Growth phenotype detection under low temperature stress
[0084] Low temperature stress can cause damage to maize leaves, as well as leaf dehydration and shrinkage. Taking wild-type maize plants as a control, the ZmbZIP68 gene knockout line constructed in Example 1 was subjected to a low temperature stress treatment test, as follows:
[0085] The 14-day-old maize seedlings (including 3-4 days of germination) grown under normal light conditions at 23°C were placed in a 4°C incubator for 3 days, and the treated ZmbZIP68CRISPR / Cas9 knockout lines D19 and D169 and the same A batch of wild type (LH244 ecotype) was placed at 23°C for 24h recovery, and then phenotype observation and picture collection were performed. The result is as figure 2 As shown, the results show that the ability of D19 and D169 to tolerate low temperature stress is significantly improved after low temperature treatme...
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