Special kit and method for detecting mRNA of gene Bmi-1 in blood plasma of patients with cervical cancer

A bmi-1mrna, bmi-1 technology, applied in the direction of fluorescence/phosphorescence, material excitation analysis, etc., can solve the problems of false negative, unsuitable screening inspection, etc., and achieve the effect of reliable result analysis

Active Publication Date: 2012-05-23
SHANDONG UNIV QILU HOSPITAL
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Problems solved by technology

The traditional cytological examination, Pap cell smear examination, has greatly reduced the mortality rate of cervical cancer, but its development has been restricted in many aspects, such as high requirements for cytological examination technicians, subjectivity in judgment, and prone to failure in obtaining materials. false negative
Colposcopy plus pathological examination is the gold standard for early cervical cancer detection, but it is not suitable for screening tests

Method used

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  • Special kit and method for detecting mRNA of gene Bmi-1 in blood plasma of patients with cervical cancer
  • Special kit and method for detecting mRNA of gene Bmi-1 in blood plasma of patients with cervical cancer
  • Special kit and method for detecting mRNA of gene Bmi-1 in blood plasma of patients with cervical cancer

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Embodiment 1

[0031] 1. The composition and preparation of the kit:

[0032]According to the gene sequences of GeneBank sequence numbers NM_005180, NM_002046 and NM_001402 as templates, the primers for detecting target gene (Bmi-1) and internal reference gene (GAPDH and EEF1A1) mRNA were designed. For the primer sequences, Tm values ​​and lengths of amplified products, see Table 1. The primer sequences all span two exons in the original gene sequence, which saves the treatment of samples with DNase and avoids the influence of genomic DNA on experimental results.

[0033] Table 1 Primer sequence, Tm value and length of amplified product

[0034]

[0035] The kit further includes PCR reaction liquid: composed of Syber Green I fluorescent dye, DNA polymerase, dNTPs, tris hydrochloride, potassium chloride and magnesium chloride.

[0036] The final concentration of the DNA polymerase in the PCR reaction solution is 100U / ml.

[0037] The final concentration of the dNTPs in the PCR reaction ...

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Abstract

The invention discloses a special kit for detecting mRNA of a gene Bmi-1 in blood plasma of patients with cervical cancer, which has the advantages of stable result, reliable analysis and the like. The special kit comprises primers of the mRNAs of the gene Bmi-1, a gene GAPDH, a gene EEF1A1, which are shown by SEQUENCE LISTING. The invention also discloses a method for detecting the expression ofthe mRNA of the gene Bmi-1 in blood plasma of the patients with cervical cancer, which has the advantages of simple, convenient and easy operation and the like. The method comprises the following step: (1) extracting total RNA of sample blood plasma; (2) performing reverse transcription-polymerase chain reaction (RT-PCR) amplification; (3) drawing a standard curve, calculating slope, and calculating amplification efficiency E according to a formula (E=10(-1 / slope)-1; and (4) calculating, namely selecting sample and calibrating sample detection holes, obtaining a sample threshold and a calculating sample threshold by using random software of real-time fluorescence quantitative PCR apparatus, obtaining a calibration initial copy number Q of the gene according to a formula (Q=(E+1)-deltaCq),and obtaining the relative expression of the target gene Bmi-1 by comparing the geometrical average of the Q values of the sample target gene and the geometrical average of the Q values of an internal reference gene.

Description

technical field [0001] The invention relates to a special kit for detecting cervical cancer plasma Bmi-1 mRNA and a method for detecting cervical cancer plasma Bmi-1 mRNA expression, belonging to the technical field of molecular biology detection. Background technique [0002] Cervical cancer is one of the most common malignant tumors in gynecology, and its incidence rate ranks second among female malignant tumors. According to statistics, there are 493,000 new cases and about 274,000 deaths worldwide every year, seriously endangering women's physical and mental health. Early diagnosis and early treatment are the key to the prevention and control of cervical cancer. If cervical cancer is found in time and properly treated, its cure rate is almost 100%. The traditional cytological examination Pap cell smear examination has greatly reduced the mortality rate of cervical cancer, but its development has been limited in many ways, such as high requirements for cytological examina...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68G01N21/64
Inventor 王传新张欣
Owner SHANDONG UNIV QILU HOSPITAL
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