The simultaneous spoligotyping, rpoB, katG, inhA loci SNPs typing is a brand-new technique to control and survey multidrug-resistant TB emergence, show to be 100% correlated to rpoB and katG hot-spot sequencing, that can be run in 3h on a single Eppendorf tube.
The technique uses the “Dual Priming Oligonucleotide” principle (DPO) and a 8 primers multiplexed PCR. It can also be run in two experiments, i.e. a 16-Plex RINT (Rifampin, Isoniazid Typing), and a 43-Plex SPOL experiment. The two modules can be purchased separately.
It consists in its full version in a 59-Plex assay (to run in a single read on Luminex 200® or BioPlex, whereas the MagPix version requires two reads).
The 59-Plex includes the detection of 43 spacers, 3 wild-type sequences on rpoB, 5 prevalent SNPs prevalent on rpoB (at position 516 (mut1), 526 (mut1, mut2), 531 (mut1, mut2), two “spanning” probes (spa_wt1, spa_wt2) that allow the indirect mutation detection of untargeted SNPs at the other positions of the rpoB Hot-Spot, and katG (wt, mut 1, mut2) and inhA (wt, mut1, mut2).
The technique was developed and validated on culture extracted DNA of MDR-TB cases and can also be used on clinical samples.
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