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Related Concept Videos

Real Time RT-PCR02:57

Real Time RT-PCR

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Real-time reverse transcription-polymerase chain reaction, or Real-time RT-PCR, is an analytical tool used to determine the expression level of target genes. The method involves converting mRNA to complementary DNA with the help of an enzyme known as reverse transcriptase, followed by the PCR amplification of the cDNA. These two processes can be performed simultaneously in a single tube or separately as a two-step reaction.
The real-time quantification of the number of amplified products is...
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Author Spotlight: Advancements and Challenges in Hepatitis B Virus Detection
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One-Step Reverse Transcriptase qPCR Method for Serum Hepatitis B Virus RNA Quantification.

Shi Liu1, Bin Zhou1, Sheng Shen1

  • 1State Key Laboratory of Organ Failure Research, Guangdong Provincial Key Laboratory of Viral Hepatitis Research, Department of Infectious Diseases, Nanfang Hospital, Southern Medical University, Guangzhou, China.

Methods in Molecular Biology (Clifton, N.J.)
|July 23, 2024
PubMed
Summary
This summary is machine-generated.

Serum hepatitis B virus (HBV) RNA offers a noninvasive method to assess intrahepatic covalently closed circular DNA (cccDNA) activity in chronic hepatitis B (CHB) patients. This protocol details HBV RNA quantification for improved clinical management without invasive biopsies.

Keywords:
DNase I digestionHBV RNAHepatitis B virusQuantificationReverse transcriptase qPCR

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A Protocol for Analyzing Hepatitis C Virus Replication
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Area of Science:

  • Virology
  • Hepatology
  • Molecular Diagnostics

Background:

  • Intrahepatic covalently closed circular DNA (cccDNA) is the template for hepatitis B virus (HBV) replication, but its direct detection requires invasive liver biopsy.
  • Chronic hepatitis B (CHB) management is hindered by the impracticality of routine intrahepatic cccDNA assessment.
  • Serum HBV RNA has emerged as a potential noninvasive biomarker for intrahepatic cccDNA activity.

Purpose of the Study:

  • To present a detailed protocol for quantifying serum HBV RNA.
  • To establish serum HBV RNA as a surrogate marker reflecting intrahepatic cccDNA activity.
  • To offer a noninvasive alternative for monitoring HBV replication in CHB patients.

Main Methods:

  • Simultaneous isolation of HBV DNA and RNA from patient serum samples.
  • DNase I digestion to eliminate contaminating HBV DNA.
  • Quantification of HBV RNA using a one-step reverse transcription quantitative PCR (RT-qPCR) assay.

Main Results:

  • The described protocol enables reliable quantification of serum HBV RNA.
  • Serum HBV RNA levels correlate with intrahepatic cccDNA activity.
  • The method provides a noninvasive approach to assess viral activity.

Conclusions:

  • Serum HBV RNA quantification is a feasible and noninvasive method for monitoring HBV cccDNA activity.
  • This protocol can aid in the clinical management of CHB patients by providing insights into viral replication.
  • The findings support the use of serum HBV RNA as a valuable biomarker in hepatitis B research and clinical practice.