Method for detecting mycobacterium tuberculosis by in-situ fluorescent loop-mediated isothermal nucleic acid amplification technology and kit

[0055] Example 1

[0056] (1) Prepare a rapid detection kit for Mycobacterium tuberculosis in situ fluorescent ring-mediated constant temperature gene amplification according to the following formula:

[0057] ——Primer mixture: The concentration of the four primers is 10pmol / μl, and the primer sequence is as follows:

[0058] Outer primer 1: 5’-CAGCACGCTAATTACCCGG-3’;

[0059] Outer primer 2: 5'-CGAGTTTGGTCATCAGCCG-3';

[0060] Inner primer 1: 5'-TGGTCGTAGTAGGTCGATGGGGAGTCGATCTGCACACAGCT-3';

[0061] Inner primer 2: 5’-Cy3-CTGCGCGATGGCGAACTCAAGGCACCGTAAACACCGTAG-3’;

[0062] ——Bst DNA polymerase: the concentration is 8U / μl;

[0063] ——Protease K: The concentration is 0.1μg / ml;

[0064] ——RNase: the concentration is 0.5mg / ml;

[0065] —— Lysozyme solution: 0.5mg / ml lysozyme, 100mM Tris-HCl (pH 8.2), 50mM EDTA;

[0066] ——LAMP reaction buffer: 10mM dNTP: 10×ThermoPol reaction buffer: 150mMMgSO 4 :5mM Betina is mixed at a volume ratio of 8:5:2:10;

[0067] ——Per liter of phosphate buffer: