Construction method of recombinant vector, constructed recombinant vector and application of recombinant vector

A recombinant vector and construction method technology, applied in the fields of molecular biology and protein, can solve the problems of tedious preparation work and double the cost, and achieve the effects of improving transfection efficiency, reducing workload and high-efficiency expression.

Pending Publication Date: 2020-07-28
ZHENGZHOU NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the two-plasmid system has problems such as cumbersome preparations before transfection, and the need for double doses of transfection reagents during transfection, which doubles the cost.

Method used

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  • Construction method of recombinant vector, constructed recombinant vector and application of recombinant vector
  • Construction method of recombinant vector, constructed recombinant vector and application of recombinant vector
  • Construction method of recombinant vector, constructed recombinant vector and application of recombinant vector

Examples

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Embodiment 1

[0116] 1. Construction of IL6 antibody 1A5 sequence eukaryotic expression vector

[0117]The phage clone 1A5 of the anti-hIL-6 protein ScFv antibody with high affinity (the nucleotide sequence of the 1A5 heavy chain is shown in SEQ ID NO: 6, and its amino acid sequence is shown in SEQ ID NO: 7; the core of 1A5-VH The nucleotide sequence is shown in SEQ ID NO: 8, and its amino acid sequence is shown in SEQ ID NO: 9; the nucleotide sequence of the 1A5 light chain is shown in SEQ ID NO: 10, and its amino acid sequence is shown in SEQ ID NO: 11 shown; the nucleotide sequence of 1A5-VL is as shown in SEQ ID NO: 12, and its amino acid sequence is as shown in SEQ ID NO: 13) as template, respectively design primer 1A5-VL upstream primer / 1A5-VL downstream primer amplification The light chain variable region 1A5-VL and the primer 1A5-VH upstream primer / 1A5-VH downstream primer amplify the DNA fragment of the heavy chain variable region 1A5-VH. PCR amplification was performed with prime...

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Abstract

The invention relates to the technical field of molecular biology and protein, in particular to a construction method of a recombinant vector, the constructed recombinant vector and an application ofthe recombinant vector. The construction method provided by the invention is simple and convenient to operate, the recombinant vector provided by the invention can express various antibody with different functions, such as IgG antibody, Fab antibody, Fv antibody, humanized ScFv-Fc antibody and mouse / human chimeric antibody and the like, and the expression efficiency of the antibody is improved.

Description

technical field [0001] The invention relates to the fields of molecular biology and protein technology, in particular to a method for constructing a recombinant vector, the constructed recombinant vector and its application. Background technique [0002] With the development of antibody engineering, phage display technology based on the ability of phage coat to express antibody fragments has emerged, that is, mRNA is extracted from splenocytes and peripheral blood lymphocytes after immunization, reverse-transcribed into cDNA, and amplified by PCR technology. Increase the heavy chain and light chain genes of the antibody, connect and clone the two into the expression vector in a certain way, and express it into a functional antibody molecule in an appropriate host cell, so as to use the antigen-antibody specific combination for screening , amplification, and then purification of antibody fragments by affinity chromatography and other means. For example, Chinese patent ZL 201...

Claims

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Application Information

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IPC IPC(8): C12N15/70C12N1/21C07K16/00C12R1/19
CPCC12N15/70C07K16/00C07K2317/24
Inventor 李闰婷陈龙欣张丽萌聂晓宁马润林王峰
Owner ZHENGZHOU NORMAL UNIV
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