Product name
HWTS-RT074A-Mycobacterium Tuberculosis Rifampicin Resistance Detection Kit (Fluorescence PCR)
Epidemiology
The kit adopts the in vitro amplification detection technology of PCR method combined with fluorescent probes. When adding a pair of primers for PCR amplification, add 4 specific wild-type fluorescent probes at the same time. Both ends of each probe are labeled with a fluorescent reporter group and a fluorescent quencher group, respectively. In the process of PCR amplification, specific primers and probes bind to the target sequence, and the formation of PCR products and the accumulation of fluorescent signals are completely synchronized by the DNA polymerase activity and 5'-3' exonuclease activity of Taq enzyme. During PCR amplification, when the target exists, four fluorescent probes can bind to the amplified fragment, but the binding efficiency of the four probes to the target is different, so the Ct value in each fluorescent channel is different. When there is a base mutation, the binding efficiency of the probe and the target is reduced, but the degree of reduction of the binding efficiency of each probe is different, and the binding efficiency of one or several probes is severely reduced or even cannot be bound to the target. If there is no base mutation in the 507-533 amino acid codon region of the rpoB gene, all four probes can bind to the target normally, there are amplification signals in all four fluorescence channels, and Ct is amplified in four fluorescence channels. There is a normal difference (△Ct) between the values. If there is a base mutation in the 507-533 amino acid codon region of the rpoB gene, the binding efficiency of the probe and the target will be reduced, and even one or several probes cannot bind to the target, the difference between the Ct values in the four fluorescence channels will become larger, even one or several fluorescence channels have no amplification signal.
Channel
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FAM
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rpoB 507-514, rpoB 513-520
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ROX
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Internal Reference
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CY5
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rpoB 520-527, rpoB 527-533
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VIC(HEX)
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38KD and IS6110
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Technical Parameters
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Storage
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≤-18℃ In dark
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Shelf-life
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9 months
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Specimen Type
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Sputum
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CV
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≤5.0%
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LoD
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rifampicin-resistant wild type: 2x103 bacteria/mL
homozygous mutant: 2x103 bacteria/mL
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Specificity
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It detects wild-type mycobacterium tuberculosis and the mutation sites of other drug resistance genes such as katG 315G>CA, InhA-15C> T, the test results show no resistance to rifampicin, which means there is no cross-reactivity.
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Applicable Instruments:
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SLAN-96P Real-Time PCR Systems
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Work Flow
Option 1.
Recommended extraction reagent: Macro & Micro-Test Viral DNA/RNA Kit( HWTS-3001, HWTS-3004-32, HWTS-3004-48) and Macro & Micro-Test Automatic Nucleic Acid Extractor(HWTS-3006).
Option 2.
Recommended extraction reagent: Nucleic Acid Extraction or Purification Reagent(YDP302) by Tiangen Biotech(Beijing) Co.,Ltd.