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Recombinant transgenic vector carrying human insulin-like growth factor-I and use thereof

A technology of transgenic vectors and growth factors, applied in the field of recombinant transgenic vectors, can solve the problems of low expression level and pollution of exogenous genes

Inactive Publication Date: 2011-11-23
SUZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Publication No. CN101270367 Chinese Patent Application Publication Instructions discloses a method for establishing a silk gland biological factory of silkworm, and a method for making oral hypoglycemic drugs by using transgenic silk gland freeze-dried powder, but the silk protein promoter is selected without a corresponding signal peptide. The expression level of exogenous genes in silk gland tissue is low, and the lyophilized powder made from silk gland tissue contains contamination of exogenous recombinant DNA, which has unpredictable risks

Method used

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  • Recombinant transgenic vector carrying human insulin-like growth factor-I and use thereof
  • Recombinant transgenic vector carrying human insulin-like growth factor-I and use thereof
  • Recombinant transgenic vector carrying human insulin-like growth factor-I and use thereof

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Experimental program
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Effect test

Embodiment 1

[0058] Embodiment one: the silk fibroin heavy chain protein gene promoter and its downstream signal peptide corresponding DNA sequence control IGF Preparation of transgenic silkworm with -Ⅰ gene

[0059] The technical operation process is as follows:

[0060] 1. IGF - Preparation of gene

[0061] According to the published cDNA sequence of human insulin-like growth factor-I gene, the DNA sequence corresponding to the human insulin-like growth factor-I active peptide (SEQ ID No. 15) was artificially synthesized according to the routine. Introduce the start codon ATG and Eco Restriction site for RV, the 3' end introduces stop codons TAA and xho I. Hin Restriction site of dIII.

[0062] SEQ ID No. 15 :TGGATATCATGggaccggagacgctctgcggggctgagctggtg gatgctcttagttcgtgtgtggagacaggggcttttatttcaacaagcccacagggtatggctccagcagtcggagggcgcctcagacaggcatcgtggatgagtgctgcttccggagctgtgatctaaggaggctggagatgtattgcgcacccctcaagcctgccaagtcagctTAAGCTTCTCGAGAT

[0063] 2. Construction of pBlue...

Embodiment 2

[0088] Example 2: Preparation and biological activity test of oral silk gland freeze-dried powder capsule

[0089] collect turn IGF- The posterior silk glands of the silkworm with the Ⅰ gene were homogenized, filtered with gauze to remove coarse impurities, and freeze-dried into powder raw materials. The raw material powder was dissolved in a ratio of 1 L of water per kg, centrifuged at 18 000 rpm for 1 hour, and the upper clear, and then freeze-dried to become fine raw material powder, and the raw material powder is packed into capsules to make oral silk gland freeze-dried powder capsules. Oral silk gland freeze-dried powder capsules have been proved by animal experiments in mice that oral administration of silkworm posterior silk gland freeze-dried powder can significantly reduce the blood sugar level of streptozotocin-induced diabetic mice by 13.43%- 34.76%. And the decline was oral dose-dependent.

Embodiment 3

[0090] Example 3: Preparation and biological activity test of oral silkworm cocoon powder capsule

[0091] After cutting the cocoons of the transgenic silkworm silkworms and removing the pupae, the cocoon shells are cut into pieces, ground into powder, and then filled into capsules to make oral cocoon powder capsules. The oral cocoon powder capsules have been proved by mouse animal experiments. After 10 days of oral administration of silk cocoon powder, the It can reduce the blood sugar level of streptozotocin-induced diabetic mice by about 49.66%. And the decline was oral dose-dependent.

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Abstract

The invention discloses a recombinant transgenic vector carrying a human insulin-like growth factor-I and a use thereof. The recombinant transgenic vector comprises an exogenous human insulin-like growth factor(IGF)-I gene, a promoter and an enhancer element; a transposons for constructing the recombinant transgenic vector is a piggyback transposons; and the promoter is fibroin heavy chain protein promoter. The recombinant transgenic vector also comprises corresponding DNA sequence of the downstream signal peptide of the fibroin heavy chain protein promoter and the sequence of a bombyx mori fibroin light-chain gene fib-LpolyA signal site. Transgenic bombyx mori producing IGF-I, which is prepared by using the recombinant transgenic vector, has a higher IGF-I expression level and can ensurethe safety of the exogenous gene and rhabdovirus.

Description

technical field [0001] The invention relates to a recombinant transgene carrier capable of expressing human insulin-like growth factor-I and a method for expressing human insulin-like growth factor-I by using a transgenic silk gland biological factory. Background technique [0002] Human insulin-like growth factor-I (IGF-I) is a single-chain polypeptide regulated by GH, consisting of 70 amino acids, with a molecular weight of 7 649 Da, and its structure is 45% similar to insulin. IGF-I has germline stability, and the structure and composition of IGF-I in all mammals are basically the same. IGF-I has a variety of physiological functions, such as promoting growth, promoting substance metabolism, and promoting cell division and proliferation. Clinically, it can be used for the treatment of diabetes, insulin resistance, obesity, hypercatabolism diseases, stagnation, renal insufficiency and nervous system diseases. [0003] At present, for the development and research of IGF-I ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/63C12N15/66C12N15/866A01K67/04A61K38/30A61P3/10
Inventor 贡成良曹广力薛仁宇郑小坚潘中华
Owner SUZHOU UNIV
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