Tegillarca granosa hemoglobin Tg-HbIIA and application thereof
A technology of hemoglobin and clam, applied in the direction of hemoglobin/myoglobin, application, peptide/protein components, etc., to achieve strong bactericidal effect and broad-spectrum bactericidal activity
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[0013] 1. Extraction and reversal of RNA from blood cells of Tegillarca granosa
[0014] Take fresh cockles, draw blood from the blood sinus of the cockles with a needle tube, take 1ml of blood into a centrifuge tube, centrifuge at 3500 rpm for one minute, remove the supernatant, save the blood cells, and extract RNA with Trizol reagent. Detect the relative absorption value of A280 and A260 with an ultraviolet spectrophotometer, and use electrophoresis to check RNA integrity. Reverse RNA with M-MLV Reverse Transcriptase Kit to obtain the first strand of cDNA.
[0015] 2. Subcloning and verification of the ORF-containing cDNA of Tg-Hb gene
[0016] Design primers based on the cDNA sequence of mature peptide of Tg-Hb gene
[0017] HbIIA-ORF-F1 5'GCAAACGAGAAGAACTAAAGCAAC 3'
[0018] HbIIA-ORF-R1 5'CCATTGATGGTTGGTCCAGATA 3'
[0019] To obtain a gene fragment encoding the mature peptide of the gene. PCR reaction program: denaturation at 94°C for 4 min, 1 cycle; denaturation at 94°C for 50 ...
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