Method for excavating testicular differential proteins of Tibetan sheep before and after sexual maturity based on DIA technology

A digging, Tibetan sheep technology, applied in instruments, measuring devices, scientific instruments, etc., can solve problems such as poor reproducibility of peptide identification

Active Publication Date: 2019-11-01
GANSU AGRI UNIV
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Problems solved by technology

In the conventional data-dependent acquisition mode, the randomness of precur

Method used

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Embodiment 1

[0030] A method for mining differential proteins in Tibetan sheep testes before and after sexual maturity based on DIA technology, comprising the following steps:

[0031] Step A. Tissue collection

[0032] In the described step A, the testicular tissues described in this patent come from male Tibetans with similar genetic background (same paternal mother) at two key developmental stages before sexual maturity (3 months old) and after sexual maturity (1 year old). Sheep: After slaughtering the sheep, take out the testis tissue, wash the external blood stains with PBS and peel off the buffy coat, cut it into small tissues of 0.5cm×0.5cm×0.5cm, and quickly freeze them in liquid nitrogen for protein samples preparation of

[0033] Step B. Protein sample preparation

[0034] Add lysate (7M urea, 2% SDS, 0.1% PMSF, 65mMDTT) to the testicular tissue, and ultrasonically lyse; after centrifuging at 4°C and 14000rpm for 30min, draw the supernatant, and use the BCA protein quantificat...

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Abstract

The invention discloses a method for excavating testicular differential proteins of Tibetan sheep before and after the sexual maturity based on a DIA (Data Independence Acquisition) technology. The method comprises the following steps: tissue collection, preparation of a protein sample, creation of a spectral database, qualitative and quantitative analysis of proteins and screening of differentialexpressed proteins. According to the method disclosed by the invention, testicular tissues of 3-month-old and 1-year-old Tibetan sheep are selected; the spectral database is analyzed and created by using a conventional DDA mass-spectrometric detection technology; and then, spectral data acquisition is performed by the DIA method, and the proteins in the sample are qualitatively and quantitativelyanalyzed, thus obtaining the differential expressed proteins in the testicles of the Tibetan sheep. The method can be used to realize quantitative screening of the differential expressed proteins inthe testicles of the Tibetan sheep, which provides a very important theoretical basis to subsequent researches on a molecular mechanism of testicular development or spermatogenesis of the Tibetan sheep under a high-altitude condition from proteomics.

Description

technical field [0001] The invention relates to the technical field of DIA quantitative methods, in particular to a method for mining differential proteins in the testes of Tibetan sheep before and after sexual maturity based on the DIA technology. Background technique [0002] As one of the three primitive sheep breeds in China, Tibetan sheep are mainly distributed in Tibet, Qinghai, Gansu, Sichuan and other places on the Qinghai-Tibet Plateau above 3000m above sea level. Sustainable development and maintaining ecological balance play a very important role. However, due to long-term grazing and lack of supplementary feeding, their production performance is low, their fecundity is low, and their sexual maturity is late, which seriously limits the reproduction of Tibetan sheep flocks, and thus restricts the economic and social development of pastoral areas. Proteins are the main participants and executors in the life activities of organisms. Therefore, it is of great signif...

Claims

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Application Information

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IPC IPC(8): G01N30/06G01N30/88
CPCG01N30/06G01N30/88
Inventor 马友记李讨讨王霞张宏豫
Owner GANSU AGRI UNIV
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