Molecular marker for detection of prostate cancer and application thereof
A molecular marker, prostate cancer technology, applied in the field of genetic engineering, can solve the problems of difficult radical cure, inconspicuous enlargement of prostate cancer, missed diagnosis or misdiagnosis, etc.
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Embodiment 1
[0034] Example 1 High-throughput sequencing screening of differentially expressed genes
[0035] 1. Sampling
[0036] From October 2012 to December 2015, 27 tissue samples were obtained from the Department of Urology at Peking Union Medical College Hospital. All samples were confirmed by pathological examination, including 8 cases of paracancerous tissue samples and 19 cases of prostate cancer samples. Store in -80°C low temperature refrigerator.
[0037] 2. Total RNA extraction from tissue samples
[0038] use Reagent (invitrogen, Cat. No. 15596-018) was used to extract sample RNA, and the experimental operation was carried out according to the product manual. The specific operation was as follows:
[0039] After collecting the samples, freeze them in liquid nitrogen. After taking them out, put the tissue into a pre-cooled mortar for grinding. After the tissue sample is powdered:
[0040] ① Add Trizol and store at room temperature for 5 minutes;
[0041] ② Add 0.2 mL of...
Embodiment 2
[0050] Example 2 RT-PCR Verification of C2CD4D Expression in Prostate Cancer Tissue and Paracancerous Tissue
[0051] 1. Materials
[0052] 34 cases of prostate cancer tissue samples and 8 cases of paracancerous tissue samples were obtained from prostate cancer samples during urological surgery in Peking Union Medical College Hospital from 2012 to 2015, and they were grouped and numbered. All samples were confirmed by pathological examination.
[0053] 2. Method
[0054] 2.1 Extract the total RNA from the tissue sample, the same as the extraction method in Example 1.
[0055] 2.2 Synthesis of cDNA by reverse transcription
[0056] use III Reverse Transcriptase (invitrogen, Cat. No. 18080-044) was used for cDNA reverse transcription, and the experimental operation was carried out according to the product manual. The specific operation was as follows:
[0057] Using a reverse transcription kit, 1 μg of total RNA was reverse-transcribed with reverse transcription buffer to ...
Embodiment 3
[0076] Example 3 Gene chip verification of C2CD4D expression in prostate cancer tissue and paracancerous tissue samples
[0077] 1, the acquisition of material, with embodiment 2.
[0078] 2, the extraction of total RNA, with the method for embodiment 1.
[0079] 3. Gene chip verification
[0080] After the total RNA was linearized and amplified, the cy3-UTP labeled and fluorescently labeled cRNAs were purified using the RNEASY Mini Kit, and the labeled cRNAs were fragmented with Amhion’s RNA Fragmentation Reagents. Human whole gene expression profile chips (4x 44K genes) from Agilent Company of the United States were used, hybridized in a chip hybridization oven at 65°C for 17 hours, then eluted, stained, and finally scanned with an Agilent DNA MicroarrayScanner scanner.
[0081]After the hybridized chip reads the data points by the chip scanner, the data is imported into the analysis software, and the mRNAs whose natural logarithm absolute value of the ratio of the two gro...
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