The reaction volume for pyrosequencing was 40 lL, containing
0.1 mol/L tris–acetate (pH 7.7), 2 mmol/L EDTA, 10 mmol/L magnesium
acetate, 0.1% BSA, 1 mmol/L dithiothreitol (DTT), 2 lmol/L
adenosine 50-phosphosulfate (APS), 0.4 mg/mL PVP, 0.4 mmol/L Dluciferin,
200 mU/mL ATP sulfurylase, 3 lg/mL luciferase, 18 U/
mL Klenow fragment, and 1.5 U/mL apyrase. Each of dNTPs was
added in the reservoir of the micro-dispenser, and pyrosequencing
reaction starts when the dispensed dNTP is complementary to the
template sequence. Ten micro litres of PCR product were used for
an assay.
The reaction volume for pyrosequencing was 40 lL, containing0.1 mol/L tris–acetate (pH 7.7), 2 mmol/L EDTA, 10 mmol/L magnesiumacetate, 0.1% BSA, 1 mmol/L dithiothreitol (DTT), 2 lmol/Ladenosine 50-phosphosulfate (APS), 0.4 mg/mL PVP, 0.4 mmol/L Dluciferin,200 mU/mL ATP sulfurylase, 3 lg/mL luciferase, 18 U/mL Klenow fragment, and 1.5 U/mL apyrase. Each of dNTPs wasadded in the reservoir of the micro-dispenser, and pyrosequencingreaction starts when the dispensed dNTP is complementary to thetemplate sequence. Ten micro litres of PCR product were used foran assay.
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