Tumor marker AQUAPORIN2 protein and application thereof
A technology of AQUAPORIN2 and tumor markers, applied in the field of tumor detection and molecular targeted therapy
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Embodiment 1
[0047] Microarray analysis of expression profile of AQP2 in different human tumor tissues and paracancerous tissues
[0048]The Tumor Genome Atlas (TCGA) project is a project jointly launched by the National Cancer Institute (NCI) and the National Human Genome Research Institute (NHGRI) in 2006. It uses large-scale sequencing-based genome analysis technology to conduct large-scale research on 36 cancers. In experiments, TCGA Genome Analysis Center (GCC) compares tumors and normal tissues to find gene mutations, amplifications or deletions associated with each cancer or subtype. To help understand the molecular mechanism of cancer and improve people's scientific understanding of the molecular basis of cancer pathogenesis.
[0049] Using the TCGA standard method to download the full gene expression profile data and clinical information of 36 tumors and their paracancerous tissues, using R language (version 3.1.1) software to filter out tumor types that do not contain AQP2 expres...
Embodiment 2
[0056] In this example, the expression of AQP2 in tumor cells and normal epithelial cells was detected by fluorescent quantitative PCR method.
[0057] 1. Materials
[0058] Head and neck squamous cell carcinoma cells SCC4 and human normal oral epithelial cells HIOEC, kidney cancer cells 786-O and human renal epithelial cells HEK293T, prostate cancer cells DU145 and human normal prostate epithelial cells RWPE-1; the above cells were purchased from ATCC cell bank in the United States.
[0059] 2. Method
[0060] 2.1 Extraction of total RNA from tumor cells and normal epithelial cells
[0061] The above six kinds of cells were respectively stored at 37°C and 5% CO 2 cultured in an incubator, cultured to a density of 90%, digested with trypsin and collected, resuspended the cells in the culture medium and counted them under a microscope, and adjusted the cell concentration to 5×10 5 cells / mL, then inoculate the cell suspension with adjusted concentration into 6-well plates, 2 ...
Embodiment 3
[0073] In this example, the expression of AQP2 protein in tumor cells and normal epithelial cells was detected by immunoblotting.
[0074] Digest and collect the six kinds of cells in Example 2 when the growth density reaches 90%, resuspend the cells in culture medium for expansion culture, then collect the cells at 80% confluence, discard the supernatant after centrifugation, and rinse with PBS Twice, discard the supernatant. Add RIPA lysate and lyse on ice for 20 min. The supernatant was collected by centrifugation at 12000 g for 10 min. Add 1X SDS sample buffer, mix well by pipetting and boil for denaturation for 5min. Total proteins were separated by 10% SDS-PAGE gel and then transferred to PVDF membrane. 5% BSA was blocked at room temperature for 2 hours, incubated with AQP2 antibody (abcam) overnight at 4°C, and washed 3 times with TBST. The secondary antibody was incubated at room temperature for 1 h, and washed 3 times with TBST. ECL ultra-sensitive chemiluminesce...
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Abstract
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Application Information
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