Encoding gene of green fluorescent protein nanobody and its preparation method and application

A green fluorescent protein and nanobody technology, applied in the field of genetic engineering, can solve the problems of high production cost, affecting the application of GFP antibody, poor stability, etc., achieving high specificity and affinity, convenient for biochemical labeling, and easy to penetrate cells. organizational effect
CN108753792BActive Publication Date: 2021-09-24SUN YAT SEN UNIV
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Patent Information

Authority / Receiving Office
CN · China
Patent Type
Patents(China)
Current Assignee / Owner
SUN YAT SEN UNIV
Publication Date
2021-09-24

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Abstract

The invention relates to four green fluorescent protein (GFP) nanobody coding genes, a preparation method and application thereof. The present invention constructs a GFP nanobody library. Using phage display technology, four nanobodies specifically binding to GFP were screened from the antibody library, named A12, E6, D5 and B9, respectively. Sequencing obtained the nucleotide sequences of these four Nanobody genes, as shown in SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3 and SEQ ID NO:4, and their corresponding amino acid sequences are shown in SEQ ID Shown in NO:5, SEQ ID NO:6, SEQ ID NO:7 and SEQ ID NO:8. The A12 gene was cloned into the transformed expression vector pADL-10b-His, and introduced into the SS320 strain; the E6, D5 and B9 genes were respectively cloned into the transformed expression vector pBAD24-Flag-His, and introduced into the TOP10 strain respectively, Thus, prokaryotic expression vectors and bacterial strains of four nanobodies were obtained. The present invention expresses and purifies four nanobodies and proves that the four GFP nanobodies can specifically bind GFP, and can be applied to the detection of GFP in basic research.
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Description

technical field

[0001] The invention relates to a coding gene of a nanobody, a preparation method and application thereof, and belongs to the field of genetic engineering, in particular to four kinds of coding genes of green fluorescent protein nanobodies, a preparation method and an application thereof. Background technique

[0002] Green fluorescent protein (GFP) is a fluorescent molecule with a size of about 27KDa, which can emit green light when excited by blue wavelength light. GFP is widely used as a protein label in the field of basic biomedical research. It is expressed by fusion with the target protein to observe the localization and molecular movement of the target protein. Therefore, GFP antibodies are in great demand in basic research.

[0003] At present, the most widely used antibody is a tetramer composed of two heavy chains and two light chains, with a size of about 150KDa, called traditional antibody. Traditional antibodies have high specificity and affini...

Claims

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