Kit capable of detecting expression quantity of BAALC (brain and acute leukemia cytoplasmic) gene mRNA (Messenger Ribose Nucleic Acid)

A kit and expression technology, applied in the field of fluorescence quantitative PCR, can solve the problems of high false positive rate of probe hybridization method, high cost of gene chip method, complicated preparation method, etc. Effects of contamination, avoiding false positives or false negatives

Inactive Publication Date: 2013-04-03
李艳 +1
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  • Application Information

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Problems solved by technology

The probe hybridization method has a high false positive rate; the gene chip method is expensive and the preparation method is cumbersome; mass spectrometry is difficult to carry out in general medical institutions; although the gene sequencing method has high sensitivity and specificity, it is expensive and cumbersome to operate. time consuming
Although the specificity and sensitivity

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  • Kit capable of detecting expression quantity of BAALC (brain and acute leukemia cytoplasmic) gene mRNA (Messenger Ribose Nucleic Acid)
  • Kit capable of detecting expression quantity of BAALC (brain and acute leukemia cytoplasmic) gene mRNA (Messenger Ribose Nucleic Acid)
  • Kit capable of detecting expression quantity of BAALC (brain and acute leukemia cytoplasmic) gene mRNA (Messenger Ribose Nucleic Acid)

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Embodiment 1

[0033] Embodiment 1. Preparation of kit of the present invention

[0034] 1. Design of specific primers and fluorescent probes

[0035] According to the gene sequence (ABL gene sequence and BAALC gene sequence are from the nucleic acid database of the National Center for Biotechnology Information, the ABL gene ID is 25, the reference sequence number is NM_005157.4; the BAALC gene ID is 79870, the reference sequence number is NC_000008 .10) Design primers and fluorescent probes specific to the above-mentioned gene sequences respectively.

[0036] 2. Prepare the components of the kit according to the composition of the following kits

[0037] The kit of the present invention consists of the following:

[0038]① RNA extraction reagent: Trizol reagent (Invitrogen, product number: 15596-026 / 100ml), add 1ml Trizol reagent to every 1ml of bone marrow tissue to quickly extract RNA from bone marrow tissue of patients with acute myeloid leukemia.

[0039] ② cDNA first-strand synthesi...

Embodiment 2

[0062] Embodiment 2. detect the expression level of BAALC gene mRNA with the kit prepared in embodiment 1

[0063] Take the detection results of bone marrow tissue samples from 30 patients with acute myeloid leukemia as an example.

[0064] The detection process of using the kit of the present invention provided in Example 1 to detect the mRNA expression of BAALC gene is as follows: first, design specific primers and fluorescent probes according to the gene sequence. Secondly, obtain bone marrow tissue samples from patients with clinical acute myeloid leukemia, quickly extract tissue RNA, and perform reverse transcription PCR to synthesize the first strand of cDNA; The standard and ABL standard were diluted to a copy number / mL of 1.0x10 3 , 1.0x10 4 , 1.0x10 5 and 1.0x10 6 , respectively to make a standard curve of the internal positive control sequence standard (such as Figure 1A with Figure 1B shown) and ABL standard standard curve (as Figure 2A with Figure 2B sho...

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Abstract

The invention discloses a kit capable of detecting expression quantity of BAALC (brain and acute leukemia cytoplasmic) gene mRNA (Messenger Ribose Nucleic Acid), belonging to the field of biotechnology. The kit comprises a detection primer and fluorescence probe, a cDNA first strand synthesis reagent, fluorogenic quantitative PCR (Polymerase Chain Reaction) mixed liquor, negative control and positive control, wherein the primers and fluorescence probes for detection include a BAALC gene primer, a reference gene ABL primer and a Taqman fluorescence probe. The BAALC gene is a sign of progenitor cell of early hematopoietic cell, and is abnormally expressed in acute and chronic patients with AML (Acute Myelocytic Leukemia), ALL (Acute Lymphocytic Leukemia) and CML (Chronic Myeloid Leukemia). According to the embodiment of the invention, the mRNA level of the BAALC gene is detected by fluorogenic quantitative PCR with high sensitivity and specificity, and the specificity and sensitivity of the detection result are both obviously improved. The kit can provide a brandnew, fast, simple and convenient gene diagnosis technology for clinically evaluating the diagnosis, prognosis and reappearance period of acute lymphocytic leukemia.

Description

technical field [0001] The invention relates to the fluorescent quantitative PCR technology in the field of biotechnology, in particular to a kit for detecting the expression level of BAALC gene mRNA. Background technique [0002] Acute myeloid leukemia (AML) is a malignant clonal disease with clinical molecular heterogeneity and is the most common type of acute leukemia. Its treatment effect is poor, and the complete remission rate after chemotherapy is 60%-70% , with a median survival of approximately 2 years. In the United States, more than 11,900 new cases occur each year. Most AML patients are middle-aged and elderly. The average age of the patients is 65 years old, and less than 10% of the cases are children. About half of adult AML patients lack chromosomal aberrations, and although these patients have a longer prognosis, only 40% survive long-term. Molecular technology can accurately distinguish the risk level of patients, so as to improve the treatment effect. Th...

Claims

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Application Information

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IPC IPC(8): C12Q1/68G01N21/64
Inventor 李艳童永清
Owner 李艳
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