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Radix isatidis protein identification method

A technology of protein identification and Radix Radix is ​​applied in the field of analytical chemistry and bioinformatics to achieve the effects of easy mastery, good repeatability and easy operation

Inactive Publication Date: 2017-09-22
NANJING UNIVERSITY OF TRADITIONAL CHINESE MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

So far, there has been no report on the proteomics research of Radix isatidis protein, so a mass spectrometry technology was invented to identify Radix isatidis protein, and at the same time, bioinformatics technology was used to study the function of Radix isatidis protein and the possible signaling pathways involved, which can fully tap Application value of Radix isatidis protein resource

Method used

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  • Radix isatidis protein identification method

Examples

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Effect test

Embodiment 1

[0040] Example 1: Mass spectrometry identification of Radix isatidis protein

[0041] 1. Experimental materials

[0042] 1.1 Instrument

[0043]Thermo Scientific EASY-nLC 1000 nanoliter liquid chromatography (Thermo Company, USA);

[0044] Millipore water purifier ultrapure water system (Millipore, Bedford, MA);

[0045] Desktop high-speed refrigerated centrifuge (Allegra X-30R type, American Beckman Coulter Company);

[0046] SDS-PAGE electrophoresis instrument (USA BIO-RAD company);

[0047] Thermo scientific EASY C18 column (2cm×100μm, 5μm);

[0048] Thermo scientific EASY C18 column (75μm×100mm, 3μm);

[0049] Fully automatic 680 microplate reader (BIO-RAD, USA);

[0050] Q-Exactive mass spectrometer (Thermo Finnigan, USA).

[0051] 1.2 Reagents and reagents

[0052] Trichloroacetic acid (TCA) (Sinopharm Chemical Reagent Co., Ltd.);

[0053] Trypsin (Shanghai Solarbio Biotechnology Co., Ltd.);

[0054] Formic acid, acetonitrile (German Merck company);

[0055] S...

Embodiment 2

[0075] Example 2: Bioinformatics analysis of Radix isatidis protein

[0076] 1 Sequence Information Extraction

[0077] A total of 1116 proteins were identified by mass spectrometry in the original data (O49485 in the protein identification results was replaced by Q8LGJ6 after the database was updated), and the protein sequence information was extracted from the NCBI database in batches (version number: release 2014_09) and saved in FASTA format (201409121009S0LYNE.fasta ).

[0078] 2 Gene Ontology (Gene Ontology, GO) functional annotation

[0079] 2.1 Sequence Alignment (BLAST)

[0080] The identified proteins were compared with the protein sequences in the NCBI nr database using the localized sequence alignment software NCBI BLAST+ (ncbi-blast-2.2.28+-win32.ext). According to the principle of similarity, the obtained functional information of homologous proteins can be used for functional annotation of target proteins.

[0081] 2.2 GO function entry extraction (Mapping) ...

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Abstract

The invention discloses a radix isatidis protein identification method. The method includes: after radix isatidis is subjected to a series of pretreatment steps including grinding, decolorizing, splitting, enzyme digestion and the like, adopting a nanoliter liquid chromatograph / mass spectrometer for separation and mass spectrometric analysis, and adopting Mascot software for database retrieval and searching; subjecting identified proteins to functional annotation and pathway annotation by bioinformatics. The method is high in protein extraction rate and large in quantity of identified protein types, and in radix isatidis proteins, 1116 types of proteins and2589 unique peptide segments are identified, and the total number of high-reliability proteins (with unique peptide sequence matching numbers larger than or equal to 2) is 742 and accounts for 66.49%. The method is suitable for identification and functional analysis of proteins in traditional Chinese medicines such as radix isatidis and has a promising application prospect.

Description

technical field [0001] The application belongs to the field of analytical chemistry and bioinformatics, in particular, it relates to a mass spectrometric identification method of Radix isatidis protein, protein function annotation and signal pathway analysis. Background technique [0002] Plant proteomics (plant proteomics) is a new discipline emerging under the background of the breakthrough of high-throughput protein analysis and identification technology and the rapid development of plant genomics. With the completion of the whole genome sequence determination of rice (Oryza sativa), Arabidopsis thaliana and poplar (Populus trichocarpa) and the deepening of genomics research, plant proteomics research has become one of the hot spots in the post-genome era. . In recent years, with the rapid development of proteomics technology, the amount of protein analysis based on various protein databases is increasing. In order to efficiently analyze the obtained data and obtain more...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N1/28G01N30/02G01N30/06G06F19/22
CPCG01N33/6848G01N1/28G01N30/02G01N30/06G01N2030/067G16B30/00
Inventor 肖平李祥段金廒陈建伟
Owner NANJING UNIVERSITY OF TRADITIONAL CHINESE MEDICINE
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