Target gene for lung cancer diagnosis and treatment
A gene and lung cancer technology, applied in the field of lncRNA markers for lung cancer diagnosis and treatment, can solve the problems of losing the best time for surgery, poor survival and prognosis of lung squamous cell carcinoma patients, etc.
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Embodiment 1
[0073] Example 1 QPCR detection of the expression of AC103718.1 in lung cancer
[0074] 1. Collect samples
[0075] 46 pairs of lung squamous cell carcinoma tissues and their corresponding paracancerous tissue samples were collected.
[0076] Inclusion criteria:
[0077] 1) The pathological diagnosis is lung squamous cell carcinoma, 2) The RNA expression level and clinicopathological information of the sample are complete.
[0078] Exclusion criteria:
[0079] 1) Suffering from other malignant tumors other than squamous cell carcinoma of the lung, 2) The case has received radiotherapy, chemotherapy and targeted drug therapy before specimen collection.
[0080] 2. RNA sample preparation and quantitative analysis
[0081] Use Trizol reagent to extract RNA from sample tissue, the steps are as follows:
[0082] Add 1mL Trizol to a glass homogenate bottle in an ultra-clean bench, weigh 50-100mg of tissue into the glass homogenate bottle, adjust the rotation speed to about 1500...
Embodiment 2
[0097] Example 2 Silencing of the AC103718.1 gene
[0098] 1. Cell culture
[0099] The lung squamous cell line H2170 cells were cultured in DMEM medium supplemented with 10% fetal bovine serum and P / S. For adherent growth of cells, place in conditions of 5% CO 2 , Cultivated in a constant temperature incubator with a humidity of 37°C.
[0100] 2. Design of siRNA
[0101] Design and synthesis of siRNA targeting AC103718.1 gene, siRNA-AC103718.1 and general negative control siRNA-NC were purchased from Shanghai Jima Gene Chemical Technology Co., Ltd. The sequence of siRNA against AC103718.1 is F: 5'-AAUUCACUCACAUUUCCAGUC-3' (SEQ ID NO.5); R: 5'-CUGGAAAUGUGAGUGAAUUCU-3' (SEQ ID NO.6) (siRNA1); sense strand: 5' -UUACAUGUAGUAACAUCUCCA-3'(SEQ ID NO.7), antisense strand: 5'-GAGAUGUUACUACAUGUAAGA-3'(SEQ ID NO.8)(siRNA2)
[0102] 3. Transfection
[0103] The day before transfection, the cells in the logarithmic growth phase were diluted to a density of 2×10 5 cells / ml, the dilu...
Embodiment 3
[0110] Example 3 Effect of AC103718.1 on lung cancer cells
[0111] 1) The effect of AC103718.1 on the proliferation of lung squamous cell carcinoma cells was detected by MTT method:
[0112] Wash the transfected cell lines with PBS, remove dead cells, digest with trypsin, prepare a single cell suspension, dilute the cell suspension, and adjust the density to 5×10 4 cells / ml, and then inoculate in 96-well culture plate, add 100 μl of cell suspension to each well, put the 96-well culture plate in 37°C, 5% CO 2 Continue culturing for 48 hours in a cell incubator, add 1×MTT to each well, continue culturing for 4 hours, discard the supernatant, add DMSO to each inner well, shake for 10 minutes, and use a microplate reader to measure each well at a wavelength of 450nm OD value of the well.
[0113] The results of MTT showed that the cell growth and proliferation rate of the transfected siRNA1 group (0.385±0.013) was significantly lower than that of the blank control group (0.598±...
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