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Volume 21, Number 3—March 2015

Noninvasive Test for Tuberculosis Detection among Primates

Tiffany M. WolfComments to Author , Lawrence Mugisha, Fernanda Miyagaki Shoyama, Melanie J. O’Malley, JoAnne L. Flynn, Benon Asiimwe, Dominic A. Travis, Randall S. Singer, and Srinand Sreevatsan
Author affiliations: Minnesota Zoological Gardens, Apple Valley, Minnesota, USA (T.M. Wolf); University of Minnesota, St. Paul, Minnesota, USA (T.M. Wolf, L. Mugisha, F.M. Shoyama, D.A. Travis, R.S. Singer, S. Sreevatsan); Conservation & Ecosystem Health Alliance, Kampala, Uganda (L. Mugisha); Makerere University, Kampala (L. Mugisha, B. Asiimwe); University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania, USA (M.J. O’Malley, J.L. Flynn)

Main Article

Table 2

Fecal IS6110 conventional and real-time PCR master mixes and reaction conditions for investigation of noninvasive tuberculosis detection in primates

PCR type Primers, 5′ → 3′ Master mix Reaction conditions
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 created: February 18, 2015
Page updated: February 18, 2015
Page reviewed: February 18, 2015
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