Gene clone of and expression Japan lamprey oral cavity gland KGD model protein possessing anti thrombotic action
An oral gland, lamprey technology, applied in genetic engineering, plant genetic improvement, peptide/protein composition, etc.
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[0034] The present invention will be further described below in conjunction with accompanying drawing.
[0035] 1. Extraction of total RNA: Trizol reagent from GIBCO BRL was used. details as follows:
[0036] (1) Take the oral gland of Japanese lamprey and quickly place it in liquid nitrogen for preservation;
[0037] (2) Weigh 0.2 g venom gland tissue, add 1 ml TRIzol reagent to prepare venom gland homogenate, and incubate at 4°C for 5 min;
[0038] (3) Add 0.2ml chloroform, close the lid tightly, shake vigorously for 15sec, and then place it on ice for 5min;
[0039] (4) 4°C, 12000xg, centrifuge for 15 minutes;
[0040](5) Transfer the upper aqueous phase to another centrifuge tube, add 0.5ml of isopropanol, and incubate on ice for 10 minutes;
[0041] (6) 4°C, 12000xg, centrifuge for 15 minutes;
[0042] (7) Discard the supernatant, add 1ml of 75% ethanol to the precipitate (containing RNA) to wash, and vortex to mix;
[0043] (8) Centrifuge at 10,000xg for 5 minutes ...
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