Bactrian camel A-FABP protein gene, recombinant protein and cloning method thereof

A recombinant protein and Bactrian camel technology, which is applied in the field of genetic engineering, can solve the problems such as the protein sequence encoded by the Bactrian camel A-FABP gene has not yet been found to be cloned, and achieves the effect of strong drought tolerance and fat deposition ability.

Inactive Publication Date: 2013-04-03
XINJIANG WANGYUAN CAMEL MILK IND
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] There are few reports on the genetic variation of the A-FABP gene in animals. So far, no reports have been found on the cloning of the A-FABP gene-encoded protein sequence of the Bactrian camel and the study on the evolution of the A-FABP gene in animals.

Method used

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  • Bactrian camel A-FABP protein gene, recombinant protein and cloning method thereof
  • Bactrian camel A-FABP protein gene, recombinant protein and cloning method thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0030] Example 1, a Bactrian camel A-FABP protein gene, the nucleotide sequence of the gene has the sequence shown in SEQ ID NO.1. The cDNA sequence of the Bactrian camel A-FABP protein gene in the present invention is by using the total RNA in the Bactrian camel liver tissue as a template, and referring to the homologous sequence design primers of the A-FABP protein genes of species such as cattle, people, and mice, The new gene sequence obtained by reverse transcription PCR. The obtained Bactrian camel A-FABP protein gene cDNA sequence is shown in SEQ ID NO.1.

[0031] According to actual needs, the above-mentioned Bactrian camel A-FABP protein gene can be further optimized or / and improved:

[0032] The nucleotide sequence of the Bactrian camel A-FABP protein gene has the sequence of 70-825 in SEQ ID NO.1. In SEQ ID NO.1, the length of the RT-PCR product is 944bp, and the electrophoresis results are shown in Figure 1, wherein the 70-72 position is the start codon ATG, the ...

Embodiment 2

[0033] Example 2, a recombinant protein of Bactrian camel A-FABP protein gene.

[0034] According to actual needs, the recombinant protein of the above-mentioned Bactrian camel A-FABP protein gene can be further optimized or / and improved:

[0035] The amino acid sequence of the recombinant protein has the sequence shown in SEQ ID NO.2. See SEQ ID NO.2 for the protein coding sequence translated from the coding sequence (CDS) in the cDNA sequence of the Bactrian camel A-FABP protein gene according to the common codons.

[0036] The recombinant protein is a polypeptide having the amino acid sequence shown in SEQ ID NO.2, or a conservative variant polypeptide thereof, or an active fragment thereof, or an active derivative thereof.

[0037] The Bactrian camel myosin gene was obtained from the Bactrian camel A-FABP protein by designing a pair of primers. The nucleotide sequence information of the primer pair is as follows:

[0038] Forward primer, SEQ ID NO.3 (forward): 5'-ATGTGCGAT...

Embodiment 3

[0040] Embodiment 3, a kind of cloning method of Bactrian camel A-FABP protein gene is characterized in that carrying out according to the following steps:

[0041] The first step, total RNA extraction

[0042] 1. Tissue separation (Isolation)

[0043] Bactrian camels were collected from Alxa League in Inner Mongolia, slaughtered quickly and liver samples were taken. The tissue samples were quickly frozen in liquid nitrogen and stored at -70°C. They were taken back to the laboratory for extraction of tissue total RNA.

[0044] 2. Total RNA isolation (Total RNA isolation)

[0045] (1) Preparation for RNA extraction

[0046] Glass products were soaked in 0.1M NaOH overnight, rinsed repeatedly with tap water, rinsed twice with distilled water, and baked at 180°C for 4 hours.

[0047] Soak the homogenizer and electrophoresis tank with 3% hydrogen peroxide for 20-30 minutes, and then rinse with 0.1% DEPC water. Since the uninactivated DEPC will affect the PCR reaction, it can be...

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Abstract

The invention relates to the technical field of genetic engineering and discloses a bactrian camel A-FABP protein gene, a recombinant protein and a cloning method thereof. A nucleotide sequence of the bactrian camel A-FABP protein gene is obtained through clone sequencing, the CDS sequence of the A-FABP gene of various species is subjected to homologous comparison, the structural features of the A-FABP gene are known and verified, and basic data are provided for a relationship for researching the A-FABP gene and the bactrian camel lipid and energy metabolism. The bactrian camel A-FABP protein is a protein with multiple biological functions and has a key regulation function in energy metabolism and fatty acid metabolism; and therefore, due to the research of A-FABP from genes, proteins and the like, a novel hope is brought to prevention and treatment of obesity, insulin-risistant symdrome, type II diabetes and other diseases, and the bactrian camel A-FABP protein gene has high application value.

Description

technical field [0001] The invention relates to the technical field of genetic engineering, and relates to a Bactrian camel A-FABP protein gene, a recombinant protein and a cloning method thereof. Background technique [0002] Adipocyte fatty acid binding protein (A-FABP), also known as FABP4 or ap2, is a member of the family of fatty acid binding proteins (FABPs). It was first discovered in adipose tissue. A-FABP is widely distributed, but is mainly expressed in adipocytes and macrophages. [0003] The human A-FABP gene is located on chromosome 8 q21, the mouse A-FABP gene is located on chromosome 3, and the pig A-FABP gene is located on chromosome 4. Compared with human, mouse and rat A-FABP gene sequences, pig A-FABP gene has 90%, 83% and 81% similarity respectively. The 4 exons of mouse A-FABP gene encode 24, 58, 34 and 16 amino acids respectively, and the 3 introns respectively encode 2629, 840 and 471 bp. The 4 exons of chicken A-FABP gene encode 24, 58, 34 and 16 ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/12C12N15/10C07K14/47
Inventor 陈钢粮
Owner XINJIANG WANGYUAN CAMEL MILK IND
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