Genetically engineered bacterium for high expression of Procambarus clarkia ATP synthase F0 subunit 8

A technology of ATP synthase and genetically engineered bacteria, applied in the field of genetic engineering, can solve the problems of decreased egg quality, shortened parental molting cycle, and increased mortality.

Inactive Publication Date: 2015-12-30
FRESHWATER FISHERIES RES CENT OF CHINESE ACAD OF FISHERY SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there will be unavoidable side effects in practice: after removal of eye stalks, it is easy to shorten the molting cycle of the parent body, increase the mortality rate, and reduce the quality of eggs and other adverse consequences

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0015] The acquisition of embodiment 1 Procambarus clarkii atp8 gene

[0016] The chemical synthesis of the whole gene was carried out according to the sequence of the atp8 gene of Procambarus clarkii (Sequence ID: AFQ31580) published in the GenBank database.

Embodiment 2

[0017] Example 2 Escherichia coli recombinant expression, purification and immunoblotting identification of Procambarus clarkii ATP synthase F0 subunit 8

[0018] Design primer atp8-1: (5′-TCAGGA GGTACC ATGCCTCAAATATCCCCTCT-3′) and atp8-2: (5′-ACTGAG AAGCTT TTATCATTTTCAAATTTTCTCAATC-3'), the 5' and 3' sides of the cDNA coding frame of the gene atp8 of Procambarus clarkii ATP synthase F0 subunit 8 were introduced into KpnI and HindIII restriction enzyme sites (underlined) respectively by PCR, The atp8 gene was inserted into the commercial expression plasmid pColdII by double enzyme digestion and molecular ligation to construct the recombinant plasmid pColdII-atp8. Then pColdII-atp8 was transformed into Escherichia coli expression strain BL21(DE3), positive transformants were screened by ampicillin antibiotic plate and verified by sequencing, and then IPTG was used to induce expression of recombinant NADH dehydrogenase subunit III protein. The medium components are: tryptone ...

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Abstract

The invention relates to a method for high expression of a Procambarus clarkia ATP synthase F0 subunit 8 protein by the aid of an Escherichia coli expression system and belongs to the technical field of genetic engineering. An atp8 gene is obtained through total gene synthesis, a recombinant expression vector pColdII-atp8 is constructed and an Escherichia coli expression strain BL21(DE3) is converted for high expression, and an important foundation is laid for subsequent deep research on a molecular mechanism of the Procambarus clarkia ovary eyestalk-ablation effect.

Description

technical field [0001] The invention relates to a genetic engineering bacterium highly expressing Procambarus clarkii ATP synthase F0 subunit 8, which belongs to the technical field of genetic engineering. Background technique [0002] How to realize the regulation of artificial ovary development with no / weak side effects has always been one of the important problems in economical shrimp and crab farming. At present, although people have explored many methods to promote the rapid maturation of shrimp and crab ovaries, such as temperature control, light control, strengthening nutritional conditions, hormone treatment, etc., eye stalk removal is still the most important method for many shrimp and crab artificial breeding to promote the rapid maturation of brood shrimp ovaries. The most common and effective method of maturation and spawning. At present, during the breeding season of shrimp and crabs, artificial removal of the unilateral eye stalk of female shrimp and crabs is ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/21C12N15/70C12N9/14C12R1/19
Inventor 水燕周鑫徐增洪沈怀舜廖元元
Owner FRESHWATER FISHERIES RES CENT OF CHINESE ACAD OF FISHERY SCI
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