Reagent kit for detecting bone marrow proliferative diseases MPLW515L mutation, special-purpose primer and probe thereof
A technology of bone marrow proliferation and kits, applied to the determination/testing of microorganisms, biochemical equipment and methods, DNA/RNA fragments, etc.
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Embodiment 1
[0040] Example 1. Design of primers and TaqMan-MGB probes for detection of myeloproliferative disease MPLW515L mutation
[0041] A pair of PCR primers were designed with Primer Express Software 2.0 software based on the MPL kinase gene (GeneLoc map region of chromosome 1, 41,576,062-45,592,722bp, GeneLoc location for GC01P043576) and its reverse complement sequence, and at the same time, according to the wild-type MPL kinase gene and MPL kinase Two TaqMan-MGB probes were designed for the reverse complement sequence of the MPLW515L mutant gene, and the 5' ends of the two TaqMan-MGB probes were respectively labeled with reporter fluorescent groups HEX and FAM, and the 3' ends were labeled with a non-luminescent quencher Group, and connected to DPI3, primers, TaqMan-MGB probe sequences are shown in Table 1:
[0042] Table 1 The sequences of primers and probes for detecting MPLW515L mutation
[0043]
[0044] The SEQ ID NO.1 in the sequence listing consists of 21 bases, which ...
Embodiment 2
[0046] Example 2. Detection of MPLW515L mutation in myeloproliferative disease
[0047] 1. Detection of MPLW515L mutation in myeloproliferative disease using primers designed according to the MPL genome gene and TaqMan-MGB probe
[0048] Use primers and TaqMan-MGB probes designed according to the MPL genome gene in Example 1 to MPDs patients, including ET, MF, unclassified MPDs, and bone marrow or peripheral blood samples (from People's Hospital) such as CML to carry out myeloproliferative For the detection of the disease MPLW515L mutation, the isolated bone marrow (NDM) of a normal donor is used as a control. The specific method is as follows:
[0049] 1) Extraction of genomic DNA from the sample to be tested
[0050] Genomic DNA was extracted from bone marrow samples of MPDs patients and normal bone marrow transplantation donors (see Table 1) under sterile conditions according to the DNA zol kit (purchased from Gibco, USA) and referring to the kit instructions. The method w...
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