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Application of human SHCBP1 gene and related product

A technology of gene and use, applied in the field of use of human SHCBP1 gene and related products, can solve problems such as functions that have not been reported yet

Active Publication Date: 2020-02-14
XUZHOU CENT HOSPITAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In thyroid cancer, the function of this gene has not been reported

Method used

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  • Application of human SHCBP1 gene and related product
  • Application of human SHCBP1 gene and related product
  • Application of human SHCBP1 gene and related product

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0126] Example 1 Preparation of RNAi lentivirus against human SHCBP1 gene

[0127] 1. Screening for effective siRNA targets against the human SHCBP1 gene

[0128] Retrieve SHCBP1 (NM_024745) gene information from Genbank; design effective siRNA targets for SHCBP1 gene. Table 1-1 lists the screened effective siRNA target sequences against the SHCBP1 gene.

[0129] Table 1-1 is targeted at the siRNA target sequence of human SHCBP1 gene

[0130] SEQ ID NO TargetSeq(5'-3') 1 TGGTGAAACCTACAATCTT

[0131] 2. Preparation of lentiviral vector

[0132] Aim at the siRNA target (take SEQ ID NO: 1 as an example) to synthesize a double-stranded DNA Oligo sequence (Table 1-2) with Age I and EcoR I restriction sites at both ends; Dicer acts on the pGCSIL-GFP vector (provided by Shanghai Jikai Gene Chemical Technology Co., Ltd.) to linearize it, and agarose gel electrophoresis identifies the digested fragment.

[0133] Table 1-2 Double-stranded DNA Oligo with sticky...

Embodiment 2

[0152] Example 2 Real-time fluorescent quantitative RT-PCR method to detect gene silencing efficiency

[0153] Human thyroid cancer TPC-1 cells and K1 cells in the logarithmic growth phase were trypsinized to make cell suspension (the number of cells was about 5×10 4 / ml) were inoculated in a 6-well plate and cultured until the cell confluency reached about 30%. According to the multiplicity of infection (MOI, TPC1:20; MOI, K1:20), an appropriate amount of the lentivirus prepared in Example 1 was added, the culture medium was replaced after 24 hours of culture, and the cells were collected after the infection time reached 5 days. Total RNA was extracted according to the instruction manual of Invitrogen's Trizol. According to the M-MLV instruction manual of Promega Company, RNA was reverse-transcribed to obtain cDNA (see Table 2-1 for the reverse transcription reaction system, react at 42°C for 1 hour, and then bathe in a water bath at 70°C for 10 minutes to inactivate the rev...

Embodiment 3

[0160] Example 3 Detection of proliferation ability of tumor cells infected with SHCBP1-siRNA lentivirus

[0161] Human thyroid cancer TPC-1 cells and K1 cells in the logarithmic growth phase were trypsinized to make cell suspension (the number of cells was about 2×10 5 / ml) were inoculated in a 6-well plate and cultured until the cell confluency reached about 30%. According to the multiplicity of infection (MOI, TPC-1:20; MOI, K1:20), add an appropriate amount of virus, and replace the medium after 16 hours of culture. After the infection time reaches 3 days, collect each experiment in the logarithmic growth phase. group of cells. The complete medium was resuspended into a cell suspension (1.5×10 4 / ml), inoculate a 96-well plate at a cell density of about 1500 cells / well. Three replicate wells per group, 100 μl per well. After laying the board, place at 37°C, 5% CO 2 Incubator cultivation. From the second day after plating, the Tecan infinite microplate instrument was ...

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Abstract

The invention belongs to the field of biological medicine research, and particularly relates to application of a human SHCBP1 gene used as a target in preparing a thyroid cancer treatment medicine orpreparing a thyroid cancer diagnosis medicine. Through the broad and deep research, the invention discovers that after the expression of the human SHCBP1 gene is down-regulated by adopting an RNAi method, the multiplication of thyroid cancer cells can be effectively inhibited, the cell apoptosis is promoted, and the growth process of thyroid cancer can be effectively controlled. The siRNA or a nucleic acid building body and a slow virus containing the siRNA can specifically inhibit a multiplication capacity of the thyroid cancer cells, inhibit a tumor formation capacity of the thyroid cancer cells in a human body, promote the thyroid cancer cell apoptosis, inhibit the cloning of the thyroid cancer cells, inhibit a thyroid cancer cell metastasis capacity, inhibit a thyroid cancer cell transfer capacity and change the period distribution of the thyroid cancer cells so as to treat the thyroid cancer, so that a new direction is opened for thyroid cancer treatment.

Description

technical field [0001] The invention belongs to the field of biomedical research, and specifically relates to the use of human SHCBP1 gene and related products. Background technique [0002] SHC SH2 domain protein 1 (SHCBP1) is an important linker protein that may be involved in MAPK-Erk, Jak-Stat signaling pathways (Genecards, https: / / www.genecards.org), and is associated with lung cancer, hepatocellular carcinoma, Malignant progression of gastric cancer, glioma, synovial sarcoma, etc. SHCBP1 is related to functional activities such as tumor cell proliferation, tumor growth and cell differentiation, but its mechanism of promoting cancer development is not very clear. [0003] By reviewing the literature, it was found that SHCBP1 is highly expressed in a variety of tumors, and its mechanisms for promoting cancer development are also different. SHCBP1 inhibits the apoptosis of lung cancer cells by regulating the expression of PTEN in lung cancer. SHCBP1 is related to the ci...

Claims

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Application Information

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IPC IPC(8): C12Q1/6886C12N15/113C12N15/867A61K45/00A61K31/713A61P35/00
CPCC12Q1/6886C12N15/113C12N15/86A61K45/00A61P35/00C12Q2600/136C12Q2600/158C12N2310/14C12N2740/15043C12N2800/107C12N2310/531Y02A50/30
Inventor 耿厚法梁军徐伟臧秀王玉腾飞孙好杰
Owner XUZHOU CENT HOSPITAL
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