Construction and screening method for recombinant human epidermal growth factor engineering bacteria
An epidermal growth factor and screening method technology, which is applied in the field of construction and screening of high-efficiency intracellular soluble expression recombinant human epidermal growth factor engineering bacteria, can solve the problems of low expression soluble rate and plasmid loss, and achieves high soluble rate , Improve the stability, simplify the effect of purification operation
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[0020] The specific process of recombinant hEGF engineering bacterium construction of the present invention is as follows:
[0021] 1. Containing the construction of kanamycin resistance gene and cspA gene functional element
[0022] 1.1 Acquisition of Kanamycin Resistance Gene Fragment
[0023] 1.1.1 Design primers, the sequence is as follows:
[0024] Primer Kan-1: 5′-ggcaaatt agatct cttttctacggggtctgacg-3' (SEQ ID NO: 1)
[0025] Primer Kan-2: 5′-aatttgcc catatg cgatttcggcctattggtta-3' (SEQ ID NO: 2)
[0026] 1.1.2 PCR reaction to obtain kanamycin resistance gene fragment
[0027] The DNA polymerase Vent used in the PCR reaction was purchased from NEB Company, and the pET42a plasmid was purchased from Merck Millipore. The PCR reaction system was as follows:
[0028]
[0029] The conditions of the PCR reaction are: denaturation: 94°C, 30 seconds; annealing: 56°C, 30 seconds; extension: 72°C, 1 minute 20 seconds; number of cycles: 30
[0030] 1.2 Acquisition of fu...
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