Kit for quantitatively detecting PML/RARalpha mRNA level

A quantitative detection and kit technology, which is applied in the field of kits for quantitative detection of PML/RARα mRNA levels, can solve problems such as incomprehension and unintuitive results of unknown samples, and achieve the effects of long storage period, low cost and cost reduction.

Inactive Publication Date: 2010-01-13
PEOPLES HOSPITAL PEKING UNIV
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Problems solved by technology

[0004] To calculate the amount of starting template, a standard curve must be used. The standard curve is made by RQ-PCR using serially diluted standards of known starting amount. The standard can be cDNA of known quality or known copy number. The former is relatively simple to make, but the result of the unknown sample obtained is not intuitive and difficult to understand; while the preparation of the plasmid standard is cumbersome, but it can be used to calculate the copy number of the unknown sample, and the result is intuitive, which is currently the most widely used plasmid. Wide range of standard types

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  • Kit for quantitatively detecting PML/RARalpha mRNA level
  • Kit for quantitatively detecting PML/RARalpha mRNA level
  • Kit for quantitatively detecting PML/RARalpha mRNA level

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Embodiment 1

[0028] Embodiment 1, the preparation of the kit of quantitative detection PML / RAR alpha mRNA level

[0029] 1. Design of primers and probes in the kit for quantitative detection of PML / RARα mRNA levels

[0030] The composition of the L-type PML / RARα real-time quantitative PCR system is as follows:

[0031] (1) Upstream primer (located in exon 6 of PML): 5'-TCTTCCTGCCCAACAGCAA-3' (sequence 1 in the sequence listing), the concentration is 0.3 μM;

[0032] (2) Downstream primer (located at RARα exon 3): 5'-GCTTGTAGATGCGGGGTAGAG-3' (sequence 2 in the sequence listing), the concentration is 0.3 μM;

[0033] (3) TaqMan probe (located in RARα exon 3): 5'-AGTGCCCAGCCTCCCTCGC-3' (sequence 3 in the sequence listing), the concentration is 0.2 μM;

[0034] (4) Universal Master Mix (purchased from American ABI Company)

[0035] The composition of the S-type PML / RARα real-time quantitative PCR system is as follows:

[0036] (1) Upstream primer (located in exon 3 of PML): 5'-CCGATGGCTTCGAC...

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Abstract

The invention discloses a kit for quantitatively detecting PML / RARalpha mRNA level. The kit comprises a standard product for manufacturing a standard curve, an internal reference gene real-time quantitative PCR system and at least one of three real-time quantitative PCR systems: L-type PML / RARalpha real-time quantitative PCR system, S-type PML / RARalpha real-time quantitative PCR system and V-type PML / RARalpha real-time quantitative PCR system. The kit can accurately, rapidly and quantitatively detect the PML / RARalpha mRNA level, is used for diagnosis of acute promyelocytic leukemia and monitoring of minimal residual diseases during treatment, and provides important molecular basis for confirmed diagnosis of clinic diseases, determination of treatment proposals, curative effect evaluation and prognosis.

Description

technical field [0001] The invention relates to a kit for quantitatively detecting the level of PML / RARα mRNA. Background technique [0002] Acute promyelocytic leukemia (APL) is a special subtype of acute myeloid leukemia. APL has specific cytogenetic abnormalities t(15;17)(q22;q12-21), resulting in promyelocytic leukemia gene (PML) on chromosome 15 and retinoic acid receptor alpha (RARα) on chromosome 17 ) forms PML / RARα fusion gene, and its translation product PML / RARα fusion protein can block the differentiation and maturation of granulocytes, and at the same time, the apoptosis of abnormal promyelocytes is insufficient, which is the main molecular mechanism of APL pathogenesis. The RARα partial break sites that form the PML / RARα fusion gene are all located on intron 2, and there are three types of break sites on PML, and the PML / RARα fusion genes are divided into three types according to the different break sites on the PML: (1 ) About 55% of APL patients are BCR1 typ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 主鸿鹄秦亚溱刘艳荣李金兰李玲娣黄晓军
Owner PEOPLES HOSPITAL PEKING UNIV
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