Molecular Genetic PML - RARA Transcrpit Qualitative Analysis - RT-PCR
Test Code
MOG50
Also Known As
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Test Parameters Included
No data available
Department
MOLECULAR GENETICS
Methodology
RT-PCR (Reverse Transcription Polymerase Chain Reaction)
This test specifically utilizes Reverse Transcription Polymerase Chain Reaction (RT-PCR). This is a qualitative assay, meaning its primary purpose is to determine the presence or absence of the PML-RARA fusion gene transcript (mRNA), rather than to precisely quantify its amount.
Principle of Qualitative RT-PCR for PML-RARA:
* RNA Extraction: Total RNA, including mRNA, is first isolated from the patient's whole blood or bone marrow sample.
* Reverse Transcription (RT): The extracted RNA is then converted into complementary DNA (cDNA) using an enzyme called reverse transcriptase. This step is essential because PCR amplifies DNA, not RNA.
* Polymerase Chain Reaction (PCR): The cDNA is then used as a template for PCR amplification. Specific primers are designed to bind only to the unique sequence formed by the fusion of the PML and RARA genes.
* Amplification: If the PML-RARA fusion transcript is present in the sample, these primers will facilitate the enzymatic creation of millions of copies of the specific fusion product.
* Detection: The amplified DNA product is typically detected through techniques like gel electrophoresis (visualizing a band of the expected size) or via probe-based methods that indicate presence.
* Internal Control: An internal control gene (e.g., ABL1, G6PD) is simultaneously amplified in the same reaction. This control confirms that the RNA extraction, reverse transcription, and PCR amplification steps were successful and that the sample is suitable for analysis, thus helping to prevent false-negative results due to technical issues or poor sample quality.
This method offers high sensitivity for detecting the PML-RARA fusion, making it a valuable tool for the initial diagnosis of Acute Promyelocytic Leukemia (APL). However, it does not provide the quantitative data needed for monitoring minimal residual disease (MRD), for which quantitative RT-PCR (RT-qPCR) is preferred.
Sample Required
Whole Blood (WB) / Bone Marrow Aspirate (BMA) collected in EDTA Vacutainer
* Approximately 3-5 mL of Whole Blood or 1-3 mL of Bone Marrow Aspirate is typically required.
* EDTA (lavender-top) tubes are the preferred anticoagulant as they are optimal for preserving nucleic acids.
* Samples must be collected and transported under specific conditions (e.g., refrigerated, not frozen) and ideally reach the laboratory within 48-72 hours of collection due to the inherent instability of RNA.
Preparation
Proper collection and handling of the sample are crucial for the accuracy and reliability of this molecular test, as RNA is inherently unstable. * No specific patient preparation is required (e.g., fa... Read more
Schedule Report
5 Days
Emergency Report
Yes
Frequently Asked Questions
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Test Description
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