Application of COCH gene and its expression product
A gene and gene chip technology, applied in the field of application of COCH gene and its expression products in the diagnosis of liver cancer, can solve the problems of unclear relationship and achieve accurate diagnosis
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Embodiment 1
[0024] Example 1 RT-PCR assay to detect the expression of COCH gene in liver cancer tissue
[0025] Reverse transcription-polymerase chain reaction (RT-PCR) was used to examine the expression of COCH gene in liver cancer tissues. RT-PCR refers to a method that combines a reverse transcription (Reverse Transcription; RT) reaction and a PCR (Polymerase Chain Reaction) reaction. RT-PCR combines RNA-templated cDNA synthesis with PCR to provide a fast and sensitive method for analyzing gene expression. RT-PCR is used to detect or quantify expression information. RT-PCR is more sensitive and easier to operate than other RNA analysis techniques including Northern blotting, RNase protection analysis, in situ hybridization and S1 nuclease analysis. The template for RT-PCR can be total RNA or poly(A) selective RNA. The reverse transcription reaction can be initiated with a reverse transcriptase, random primers, oligo(dT), or gene-specific primers. RT-PCR can be performed in a one-st...
Embodiment 2
[0049] Example 2 Realtime-PCR assay to detect the expression of COCH gene in liver cancer tissue
[0050] Relative quantitative method was used to detect the expression difference of STC2 gene in liver cancer samples (Thermal Cycler Dice TM RealTime System TP800, Takara; Premix Ex Taq TM , Takara).
[0051] The fluorescent quantitative PGR (real-time PGR) reaction system is as follows: total volume 20 μL, SYBR Premix Ex Taq 10 μL, upstream and downstream primers (10 μmol / L) 0.4 μL each, cDNA 1 μL, ddH2O 8.2 μL, mix the reagents, centrifuge and put The amplification reaction was carried out in a PGR instrument. The reaction conditions were denaturation at 95°C for 5 seconds, annealing and extension at 68°C for 30 seconds, 40 cycles. The instrument was operated according to the manufacturer's instructions. The housekeeping gene β-actin was used as an internal reference.
[0052] The primers designed in this example are as follows:
[0053] COCH-F: 5'-ACAGGAAAAGCCTTGAAGCA...
Embodiment 3
[0058] Example 3 PGR experiment detects the expression of COCH gene in normal human tissue
[0059] The expression of COCH gene in normal adult tissues was detected by conventional PGR method. Experimental results show that COCH is expressed in embryonic liver but not in mature liver (see image 3 ), further indicating that the expression of COCH gene can be used for accurate diagnosis of liver cancer.
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