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Fusion expressed product of vascular inhibine and endostatin in colibacillus and its preparing method

A technology of endostatin and angiostatin, applied in the field of genetic engineering technology, can solve problems such as damage, toxic side effects, and inevitable normal cells

Active Publication Date: 2007-06-27
TIANJIN JINSITAN BIOLOGICAL TECH
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Problems solved by technology

Traditional drug therapy is mainly aimed at abnormally proliferating tumor cells, but while traditional antineoplastic drugs kill tumor cells, normal cells are also inevitably damaged, causing serious side effects

Method used

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  • Fusion expressed product of vascular inhibine and endostatin in colibacillus and its preparing method
  • Fusion expressed product of vascular inhibine and endostatin in colibacillus and its preparing method
  • Fusion expressed product of vascular inhibine and endostatin in colibacillus and its preparing method

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Embodiment Construction

[0034] The present invention will be further described below in conjunction with the drawings and embodiments:

[0035] 1. RNA extraction:

[0036] Add 1ml TRIZOL to 50g-100g liver tissue. Centrifuge at 12000rpm at 4°C for 10 minutes to collect the supernatant. Incubate the sample at room temperature for 5 minutes, add 0.2ml of chloroform per 1ml of TRIZOL, shake for 15 seconds, incubate at room temperature for 3 minutes, centrifuge at 4°C and 12000rpm for 15 minutes to collect the supernatant. Add 0.5ml isopropanol per 1ml TRIZOL to incubate at room temperature for 10 minutes, and centrifuge at 4°C at 12000rpm for 10 minutes to discard the supernatant.

[0037] Wash the pellet with 75% ethanol, mix the pellet, and centrifuge at 4°C at 8000 rpm for 5 minutes. Dry the precipitate, dissolve it in DEPC water and store at -20°C.

[0038] 2. RNA reverse transcription

[0039] Follow Invitrogen's reverse transcription kit instructions. Take 3μl total liver RNA, 2μl downstream primers, 1...

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Abstract

The present invention discloses a kind of fusion expressed product of vascular inhibine and endosatin in colibacillus and its preparation process, and belongs to the field of genetic engineering and peptide-containing medicine preparation. Through extraction of RNA from liver tissue of animal, RNA reverse transcription, PCR amplification of ES and AS segment and recovering ES and AS segment, and the connection reaction of introducing NdeI, NheI and XholI cleavage site to the primer via the SacI cleavage site in pGEM-T Easy vector to connect vascular inhibine and endosatin, fusion expressed product of vascular inhibine and endosatin in colibacillus is obtained. The fusion expressed product has not only the functions of both vascular inhibine and endosatin but also the synergistic effect of antagonizing angiogenesis and tumor.

Description

Technical field [0001] The present invention relates to genetic engineering technology, or pharmaceutical preparations containing peptides, in particular to a fusion expression product of angiostatin and endostatin in Escherichia coli and a preparation method thereof. Background technique [0002] Tumor is one of the most difficult diseases in the world today. Traditional drug therapy mainly targets abnormally proliferating tumor cells. However, while traditional anti-tumor drugs kill tumor cells, they inevitably damage normal cells, causing serious side effects. New anti-tumor genetic engineering drugs, the endogenous vascular inhibitory factors angiostatin (AS) and endostatin (ES), because of their specific action and no toxic side effects, have received more and more attention in tumor treatment. Recent studies have shown that the combination of the two has synergistic anti-angiogenesis and anti-tumor effects. [0003] Chinese patent 1324818 discloses a "method for producing e...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/62C12N1/21C12N15/12C12N15/09C12P21/02C07K19/00
Inventor 王卫星
Owner TIANJIN JINSITAN BIOLOGICAL TECH
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