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Novel vector for raising content of polyunsaturated fatty acids in animal body

A technology of unsaturated fatty acids and animal bodies, applied in the field of construction of new expression vectors, to achieve the effect of increasing the content and increasing the content

Inactive Publication Date: 2011-10-05
INST OF ANIMAL SCI OF CHINESE ACAD OF AGRI SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] Linoleic acid and linolenic acid are essential fatty acids in mammals, but mammals cannot synthesize them by themselves due to the lack of Δ12 and omega-3 fatty acid dehydrogenase, so they can only be ingested from food

Method used

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  • Novel vector for raising content of polyunsaturated fatty acids in animal body
  • Novel vector for raising content of polyunsaturated fatty acids in animal body
  • Novel vector for raising content of polyunsaturated fatty acids in animal body

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Embodiment 1

[0025] Construction and functional evaluation of embodiment 1 eukaryotic expression vector

[0026] 1. Research Methods

[0027] 1. Vector Construction

[0028] 1) Analyze the complete gene sequences of sFat-1 and Fat-2 sequences, and check whether the genes contain complex secondary structures and repetitive sequences. Design a synthesis scheme according to the specific conditions of the sequence, first design and synthesize a single-stranded oligo, and use PCR to splice the synthesized oligo into a complete gene.

[0029] 2) Load the synthesized sFat-1 gene into the pEASY-Blunt Simple vector and transform it into a competent cell Trans1-T1 Phage Resistant Cell, and verify whether the gene sequence in the recombinant clone meets the requirements by sequencing.

[0030] 3) KpnI / NotI double-enzyme digestion of the vector containing the target fragment synthesized by the whole gene to obtain a 1398bp target fragment, and KpnI / NotI double-enzyme digestion of the pcDNA3.1 vector...

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Abstract

The invention discloses a novel vector for raising the content of polyunsaturated fatty acids in an animal body. The expression vector provided by the invention can simultaneously transcribe and express omega-3 fatty acid dehydrogenase and delta12 fatty acid dehydrogenase, and the vector sequence contains gene sequences of Sequence NO.1 and Sequence NO.2. In addition, the sequences of the enhancer SP163 and IRES are added between the sFat and Fat2 genes, and the vector provided by the invention can highly express two types of dehydrogenase at the same time. The vector provided by the invention is also transfected into the CHO cells, followed by the screening of antibiotics to obtain a stable cell line for highly expressing the sFat and Fat2 genes with the gene expression amount being 105 times that of untransfected cells. The invention also discloses a concrete method for raising offspring animals having high content of polyunsaturated fatty acids by using the vector. The vector provided by the invention lays a foundation for producing animal products having high content of omega-3 polyunsaturated fatty acids and improving the quality of meat products or dairy products by producing transgenic livestock.

Description

technical field [0001] The invention relates to the construction of a novel expression vector. Specifically, the sequences of genes sFat-1 and Fat-2 were cloned from the nematode Caenorhabditis elegans and codon optimized. The invention constructs a high expression vector pcDNA3.1-sFat-SP163-IRES-Fat2, which can be applied to increase the content of ω-3 polyunsaturated fatty acids in animals. Background technique [0002] Polyunsaturated fatty acids (PUFAs) refer to fatty acids with more than sixteen carbon atoms and more than two double bonds, and ω-3PUFAs and ω-6PUFAs are the two most important types. α-linolenic acid (α-linolenic acid, αLA), eicosapentaenoic acid (EPA), docosapentaenoic acid (DPA), docosahexaenoic acid (DHA) All belong to ω-3PUFAs; while linoleic acid (linoleic acid, LA), γ-linolenic acid (γ-lenolenic acid, γ-LA), arachidonic acid (arochidonic acid, AA) and so on belong to ω-6 PUFAs. More and more studies have proved [Chen W X, Chen D G, Cheng X F, etc...

Claims

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Application Information

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IPC IPC(8): C12N15/85A01K67/027
Inventor 苗向阳
Owner INST OF ANIMAL SCI OF CHINESE ACAD OF AGRI SCI
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