A yess-based method for analyzing protein interactions in prokaryotes

A protein interaction, prokaryotic technology, applied in the field of protein interaction analysis in prokaryotes based on YESS, can solve problems such as false positives and false negatives, inappropriate proteins, and inability to quantify the strength of protein interaction relationships.

Active Publication Date: 2021-04-02
武汉睿嘉康生物科技有限公司
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Problems solved by technology

[0006] In the existing general technology, the false positives and false negatives caused by technical problems or species homology problems are too high
[0007] Existing techniques fail to quantify the relationship strength of protein interactions in prokaryotes
[0008] Existing techniques, capable of high-throughput methods, are not applicable to all types of proteins

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  • A yess-based method for analyzing protein interactions in prokaryotes
  • A yess-based method for analyzing protein interactions in prokaryotes
  • A yess-based method for analyzing protein interactions in prokaryotes

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Embodiment Construction

[0081] In order to make the object, technical solution and advantages of the present invention more clear, the present invention will be further described in detail below in conjunction with the examples. It should be understood that the specific embodiments described here are only used to explain the present invention, not to limit the present invention.

[0082] In the existing yeast two-hybrid technology, the false negatives and false positives are too high in the actual application process; the transformation efficiency is low; the two proteins are on different vectors, and their expression levels are affected by the copy number of the vector; at the same time, because the reaction occurs in the nucleus , so it does not apply to all proteins.

[0083] The pull-down technique requires purified proteins, cannot persist transient protein interactions and has false positives.

[0084] The host of E. coli hybridization technology is prokaryote, which will cause many false posi...

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Abstract

The invention belongs to the technical field of gene engineering, and discloses a method for analyzing protein interaction in prokaryotes based on a Yeast Endoplasmic reticulum Sequestration System (YESS). The method includes the steps: constructing an expression plasmid pESD-PPI-GFP-mCherry capable of performing Golden Gate assembling, with gfp and mCherry fluorescent genes, based on pESD-PPI plasmid; selecting 5 mutually interacted protein pairs from the existing data of Zymomonas mobilis on a UniProt database and the protein pairs which are not reported to have interaction as positive and negative contrasts for studying protein interactions of proteins in Zymomonas mobilis; and quantifying the analysis results. The method for analyzing protein interactions in prokaryotes can achieve high-throughput and quantitative study on protein interaction in Zymomonas mobilis. The method for analyzing protein interactions in prokaryotes based on YESS has many advantages of being simple, being easy to operate, being highly efficient, being sensitive and the like.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and in particular relates to a method for analyzing protein interaction in prokaryotes based on YESS. Background technique [0002] At present, the existing technologies commonly used in the industry are as follows: [0003] The study of protein-protein interactions is of great significance in many biological fields, and it is very important for the analysis of various metabolic processes, regulatory networks and molecular mechanisms of energy conversion in cells. [0004] At present, methods for verifying protein-protein interactions mainly include several classic experimental techniques: yeast two-hybrid technique, E. coli hybridization technique, pull-down technique and so on. Yeast two-hybrid technology is characterized by sensitivity, high efficiency, and high-throughput automation. However, there are many shortcomings in the actual application process, such as too high false n...

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G16B20/00G16B25/00C12N15/66C12N15/70C12N15/81
Inventor 杨世辉杨青唐莹易犁
Owner 武汉睿嘉康生物科技有限公司
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