Recombinant plasmid for curing prostate gland cancer and melanoma

A melanoma, recombinant plasmid technology, applied in gene therapy, recombinant DNA technology, introduction of foreign genetic material using vectors, etc., can solve the problem of no local injection of recombinant plasmids for prostate cancer and melanoma.

Inactive Publication Date: 2006-02-08
JILIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The invention provides a recombinant plasmid for treating prostate cancer and melanoma to solve the problem that there is no local injection of recombinant plasmid for treating prostate cancer and melanoma at present

Method used

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  • Recombinant plasmid for curing prostate gland cancer and melanoma
  • Recombinant plasmid for curing prostate gland cancer and melanoma
  • Recombinant plasmid for curing prostate gland cancer and melanoma

Examples

Experimental program
Comparison scheme
Effect test

experiment example

[0026] Experimental example The recombinant plasmid PcDNA3.1(+)-U6stat3 siRNA-GRIM19 of the present invention inhibits the growth of prostate cancer in nude mice and melanoma in C57 mice

[0027] 1 In vivo tumor inhibition test in tumor-bearing nude mice

[0028] 15 nude mice, each 2×10 6 / 100ul PC3 cells were inoculated to the left ventral wall, measured with a vernier caliper every day, and the volume was according to the formula m 1 2 × m 2 ×0.5236 calculation, where m 1 is the short axis distance, m 2 is the long-axis distance, and when the tumor volume grows to a certain size, the animals are divided into four groups, 1, blank saline control group, 2 negative (siRNA expression vector constructed with 19 base sequences that are not homologous to all sequences in the human body) ) control group; 3pSilencerU6-stat3siRNA group; 4PcDNA3.1-siRNAstat3-GRIM19 recombinant plasmid group. The dose of plasmid injection was 20 μg / mouse (repeated administration at intervals of 7 ...

Embodiment

[0031] Embodiment: invention recombinant plasmid construction process

[0032](1) Construction of pSilencer U6-Stat3 siRNA vector: The pSilencer U6 plasmid was purchased from Ambion Corporation in the United States. According to the known human Stat3 mRNA sequence NM003150 in Gene Bank, a suitable target site (2143-2162) was found to synthesize a DNA template encoding siRNA , after linearizing pSilencerU6 with BamH I and Hind III, it was ligated under the action of T4 ligase;

[0033] (2) Construction of pCXN2mycA-GRIM19 expression vector: The coding sequence of GRIM19 was fished out by RT-PCR method, and the expression vector pCXN2mycA was purchased from Invitrogen Company of the United States. After linearization with KpnI and EcoRI, the two were combined with T4 ligase connect;

[0034] (3), use (1) as a template by PCR method to capture the coding sequence encoding U6 promoter and Stat3 siRNA; the PcDNA3.1(+) expression vector was purchased from Invitrogen Company in the ...

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Abstract

The invention relates to a recombinant plasmid for treating prostatic csarcinoma and melanoma, wherein the GRIM19 expression sequences are located under the CMV promotor of the vector PcDNA3.1(+) and linked by Kpn I and EcoR cleavages, the U6 promotor-siRNA-stat3 specific sequence is located in front of the PcDNA3.1(+)CMV promotor, and linked by Bgl II and NruI cleavages.

Description

technical field [0001] The invention belongs to the field of biotechnology, specifically a recombinant plasmid, and relates to the application of RNAi technology to inhibit the overexpression of Stat3 in prostate cancer and melanoma to achieve the purpose of treating tumors. GRIM19 can obviously enhance its inhibitory effect on tumors. Background technique [0002] The excessive activation and expression of the signal transducer and transcriptional activator Stat3 can lead to unlimited cell growth, resistance to apoptosis, and canceration. At present, the overexpression of Stat3 has been found in many kinds of malignant tumors, including prostate cancer and melanoma. Therefore, using Stat3 as a target for tumor treatment will bring new hope to the treatment of tumors. [0003] GRIM19 is a gene expressed in normal tissues. Studies have found that its expression is turned off when carcinogenesis occurs, and introducing its expression plasmid into cancer cells can induce apopto...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/63A61K48/00A61P35/00
Inventor 赵雪俭徐德启高丽芳张灵
Owner JILIN UNIV
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