Yeast genetically engineered strain expressing hrp gene and constructing method thereof
A technology of genetically engineered strains and construction methods, applied in the field of yeast genetically engineered strains and their construction, can solve the problems of small harvest, complicated process, and inability to meet market needs, etc.
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[0108] Construction method of genetic engineering strain HrpZ
[0109] 1. Genomic DNA extraction and hrpZ gene isolation and cloning
[0110] 1. Genomic DNA Extraction
[0111] A strain of Pseudomonas syringae was isolated and identified from the soil, and the genomic DNA was extracted according to the instructions of the Genomic DNA Extraction Kit from QIAGEN Company.
[0112] 2. Isolation of hrpZ gene
[0113] The hrpZ gene was amplified and isolated by PCR using the genomic DNA of Pseudomonas syringae as a template, and the PCR primer sequences were as follows:
[0114] Forward: 5'-ATGCAGAGTCTCAGTCTTAACAGCA-3'
[0115] Reverse: 5'-AGTTCTCCGTCAGGCGGTTGTC-3'
[0116] PCR reaction conditions:
[0117] Pre-denaturation: 94°C for 1 minute
[0118] Denaturation: 92°C for 45 seconds
[0119] Refolding: 66°C for 45 seconds
[0120] Extension: 68°C for 2 minutes
[0121] Cycles: 30
[0122] Termination of extension: 68°C for 20 minutes
[0123] The PCR product was detecte...
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