Application of biomarker in breast cancer
A breast cancer and drug technology, applied in the field of biomedicine, can solve the problems of genome regulation and specific mechanism are not very clear
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Embodiment 1
[0050] Example 1 Screening for gene markers associated with breast cancer
[0051] 1. Sample collection
[0052] The cancer tissue and corresponding normal tissue samples (5 cm from the tumor edge) of 4 cases of Luminal B breast cancer were collected for high-throughput sequencing. All patients did not receive chemotherapy, radiotherapy and endocrine therapy before operation, and all patients were informed Agree, all the above specimens were obtained with the consent of the organizational ethics committee, and the patient information is shown in Table 1.
[0053] Table 1 Sample Information
[0054]
[0055] 2. RNA sample preparation and quality analysis
[0056] Extraction of tissue total RNA using TRIZOL method
[0057] 1) Mince the tissue with scissors, add 1ml Trizol, shake on the shaker for 1min; place at room temperature for 10min to completely decompose the ribosomes.
[0058] 2) Add 200 μl of chloroform (chloroform), close the cap tightly, shake vigorously for 15...
Embodiment 2
[0088] Example 2 QPCR sequencing to verify the differential expression of LINC01781 and LINC02014 genes
[0089] 1. Large-scale QPCR verification of the differential expression of LINC01781 and LINC02014 genes was performed on cancer tissue samples and normal tissue samples collected from 25 cases of luminal B breast cancer patients collected according to the collection method in Example 1.
[0090] 2. RNA extraction
[0091] Tissue RNA was extracted by the Trizol method, see Example 1 for specific steps.
[0092] 3. Reverse transcription: the reverse transcription kit (Takara code: DRR047A) of TAKARA Company was used for operation.
[0093] 1) Removal of genomic DNA
[0094] Add 5×gDNA Eraser Bμffer 2.0μl, gDNA Eraser 1.0μl, total RNA 1μg, add RNase Free ddH in the test tube 2 O to bring the total volume to 10 μl and heat in a water bath at 42°C for 2min.
[0095] 2) Reverse transcription reaction
[0096] Will Buffer 2 4.0μl, RT Enzyme Mix I 1.0 μl, RTPrimer Mix 1.0...
Embodiment 3
[0117] Example 3 Expression of LINC01781 and LINC02014 in breast cancer cell lines
[0118] 1. Cell culture
[0119] The BT474 cell line of Luminal B breast cancer was cultured in DMEM medium containing 10% fetal bovine serum (Gibco Company) in 5% CO 2 , cultured in a constant temperature incubator at 37°C.
[0120] 2. Transfection
[0121] The general siRNA-NC, siRNA-LINC01781 and siRNA-LINC02014 used in this application were purchased from Shanghai Jima Pharmaceutical Technology Co., Ltd., and the siRNA sequences for silencing LINC01781 and LINC02014 are shown as siRNA1 (LINC01781) and siRNA2 (LINC02014), respectively.
[0122] As follows.
[0123] Sequence of siRNA1:
[0124] The sense strand is 5'-UUCUGUUACUGGUAUCAGCCA-3' (SEQ ID NO.7)
[0125] The antisense strand is 5'-GCUGAUACCAGUAACAGAAAA-3' (SEQ ID NO.8)
[0126] Sequence of siRNA2:
[0127] The sense strand is 5'-UAUACACUAGAUUUCCAAGCC-3' (SEQ ID NO.9)
[0128] The antisense strand is 5'-CUUGGAAAUCUAGUGUAUAUU-...
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