Volume 21, Number 3—March 2015
Noninvasive Test for Tuberculosis Detection among Primates
|PCR type||Primers, 5′ → 3′||Master mix||Reaction conditions|
||Forward: TTCAGGTCGAGTACGCCTTC Reverse: CGAACTCAAGGAGCACATCA
||12.5 μL HotStarTaq Master Mix:* 8 μL RNase-free water,* 0.4 μM of each primer, 1.25 μL DMSO, 0.25 μL 1% BSA, 1 μL DNA template. Total volume 25 μL
||95°C for 15 min/DNA polymerase activation; 40 cycles: 94°C for 30 s/denaturation, 56°C for 30 s/annealing, 72°C for 1 min/extension. Termination at 72°C for 10 min
|Real-time||Forward: AGAAGGCGTACTCGACCTGA Reverse: CCGGATCGATGTGTACTGAG||LightCycler 480 Probes Master:† 0.2 mM of each primer, 0.2 mM of the FAM-labeled IS6110 probe,† 5 μL DNA template. Total volume 25 μL||95°C for 5 min; 45 cycles: 95°C for 10 s/denaturation; 50°C for 30 s/annealing; 72°C for 1 s/extension. Termination at 65°C–95°C at 2.2°C/s/melting curve analysis|
*QIAGEN, Inc., Valencia, CA, USA.
†Roche, Indianapolis, IN, USA
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Page updated: February 18, 2015
Page reviewed: February 18, 2015
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