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Spider silk protein gene designed and synthesized by plant preference codon and its application

A spider silk protein and codon-preferred technology, which is applied to spider silk protein genes designed and synthesized according to plant-preferred codons and their application fields, and can solve problems such as skin allergies

Inactive Publication Date: 2003-01-15
SHANGHAI JIAO TONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In addition, even if a large amount of spider silk protein is obtained by recombining animal cells, the skin allergies of some people are caused by various synthetic chemical components remaining in the process

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0132] According to the nucleic acid sequence of the spider silk protein spidroin, the compound spidroin gene-I (Super Fiber Strength Gene, SFSG-I) is designed and synthesized according to the plant preferred codon comparison table, with a full length of 2038bp (including Linker), its nucleic acid sequence is SEQ ID NO.1, wherein the open reading frame is located at 25-2038 nucleotides. According to the deduced amino acid sequence of the full-length SFSG-I, it has a total of 663 amino acid residues, a molecular weight of 51.2KD, and a pI of 7.3. See SEQ ID NO.2 for the detailed sequence.

[0133] In order to confirm that the synthesized gene has no gene function change with the published spider silk protein gene, the present invention conducts information and homology analysis on the synthesized gene.

[0134] The full-length sequence of SFSG-I and its encoded protein were compared with the nucleotide and protein in the Non-redundant+GenBank+EMBL+DDBJ+PDB and Non-redundant Gen...

Embodiment 2

[0151] According to the nucleic acid sequence of the spider silk protein fibroin-3 (ADF-3), the present invention designs and synthesizes the compound spider silk protein gene-II (super fiber strength gene SFSG-II) according to the comparison table of plant preferred codons. The full length is 2000bp (including linker), its nucleic acid sequence is SEQ ID NO.3, wherein the open reading frame is located at nucleotides 25-2000. According to the deduced amino acid sequence of the full-length SFSG-II, it has a total of 654 amino acid residues, a molecular weight of 60KD, and a pI of 6.8. See SEQ ID NO.4 for the detailed sequence.

[0152] The full-length sequence of SFSG-II and its encoded protein were carried out by BLAST program in the Non-redundant+GenBank+EMBL+DDBJ+PDB and Non-redundant GenBank CDS translations+PDB+SwissProt+Superdate+PIR databases for nucleotide and protein synchronization. According to the source search, it was found that it has a certain homology with the w...

Embodiment 3

[0169] The gene (SFSG-I or SFSG-II) synthesized in Example 1 and Example 2 was produced and expressed in cotton. Construction of Agrobacterium expression vector

[0170] Under the premise of ensuring that the reading frame is correct, inoculate Escherichia coli containing the PBI121 plasmid vector of the synthetic gene (SFSG-I or SFSG-II) into LB medium containing ampicillin, and cultivate overnight on a constant temperature shaker at 37°C . Plasmids were extracted from overnight bacteria following the protocol provided by Molecular Cloning.

[0171] Then transfer it into the plant expression vector of Agrobacterium (such as EHA105 or LBA4404), and then use the Agrobacterium to transform cotton. Agrobacterium transformation method

[0172] Pick a single colony and inoculate it in 5ml YEB liquid medium, shake overnight at 28°C and 250rpm.

[0173] Take 2ml of bacterial liquid and transfer it into 50ml of YEB medium to continue culturing until OD600 reaches 0.5.

[0174] Tr...

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Abstract

The present invention relates to a spider silk crystal protein gene synthesized by plant preferance codon, its expression carrier, the plant cells transformed from it, and the transgenic plant and its descendant. Said gene can be effectively expressed in fibrous plant and the obtained protein moleculae can be combined with the cellulose of plant, so improving the strength of plant fibres.

Description

Technical field: [0001] The present invention relates to a spider silk protein gene designed and synthesized according to plant preferred codons and its application. The composite spider silk protein with the same strength as natural spider silk protein and its coding sequence are efficiently expressed in fiber plants, and the obtained The combination of protein molecules and plant cellulose has the characteristic of significantly enhancing the strength of plant fibers, and the formed composite plant fibers can be used in the production of textile industry. The invention relates to technical fields such as molecular biology and genetic engineering. Background technique: [0002] Long-term studies have found that spiders can produce webs and draglines with extremely high tensile strength. Since spider silk has extremely high tensile strength (several times higher than silk and 5 times stronger than steel), it is stronger than Kevlar (9.5×10 4 / 3×10 4 JKg -1 ) is several t...

Claims

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Application Information

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IPC IPC(8): C12N15/12C12N15/82
Inventor 唐克轩左开井孙小芬
Owner SHANGHAI JIAO TONG UNIV
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