Soil protein extraction and intracellular protein separation method

Abstract

The invention provides a high-efficiency soil protein extraction method, comprising the following steps: adding soil in a pre-cooling soil protein extraction buffer solution and blending and oscillating the mixture in a 4 DEG C shaking table over night; performing drawing and filtering of the shaken mixture in a buchner funnel using a crude filter paper; processing the drawing and filtering liquid by macroporous resin (D101), filtering the processed liquid through a 0.22 Mum microporous membrane, the filtered liquid being the extracellular protein mother liquid; processing the cells on the filter membrane by cleaning, crushing, centrifugalizing and vacuum drying to obtain the dried powder, namely intracellular soil protein sediment. The extracellular protein mother liquid is processed by centrifuging, separating and vacuum drying to obtain the intracellular soil protein dried powder; degrading the intracellular soil protein dried powder until the degraded intracellular soil protein dried powder is clearly separated on 10% polyacrylamide gel SDS-PAG electrophoresis, the method has high separation efficiency, microbial yield is 37.5%, which is higher than the yield of the traditional method 0.1-1%, simple operation flow, good SDS-PAGE resolution and good mass spectrum identification effect.

Description

technical field [0001] The invention relates to a method for extracting soil protein and separating intracellular protein in the field of biotechnology. technical background [0002] Metaproteomics is a new concept and research technology that has emerged in recent years. In addition to the study of microbial populations in the environment, this technology can also conduct in-depth analysis of microbial communities: through the study of metaproteomics, the composition of microbial communities and their functions can be compared. to better understand the microbiome. Although few metaproteomics studies have been carried out, metaproteomics has initially demonstrated its powerful functions in the research fields of functional gene expression in special extreme environments, the development of special functional proteins, and the cycle of ecological elements. [0003] For the first time, Wilmes et al. took activated sludge with biological phosphorus removal function as the rese...

AI Technical Summary

Problems solved by technology

However, there is still no general protein extraction method applied to environmental samples, especially from complex environmental samples such as seawater and soil, it is still very difficult

Existing studies such as AMD biofilm and activated sludge, these samples come from extreme environments with low microbial diversity. For seawater, soil and other samples with complex environments and high microbial diversity, the traditional plate method can only provide Less than 1% of the microbial information, and the method established in 1993 (O.A.Ogunseitan, 1993), which is now widely used by metaproteomics researchers, we have repeated and failed to obtain ideal results, indicating that the method lacks good versatility Therefore, the development of efficient and general protein extraction methods is very important for the research and application of metaproteomics