1.Purpose of the experiment:
Purification and identification of SFRP1 recombinant protein;
*The function of SFRP1 recombinant protein in regulating the Wnt signaling pathway.
2.SFRP1 Basic Information:
SFRP1 (Secreted Frizzled Related Protein 1) is a secreted protein encoded by the human gene SFRP1.
SFRP1 is an important regulator in the Wnt signaling pathway, which can regulate the transmission of Wnt signaling by binding to Wnt protein.
SFRP1 exerts its function both inside and outside the cell, and is closely related to the occurrence and development of cells, as well as muscle development.
Catalog No. PA1000-5532
3.Protocol Procedure:
Step 1: Expression and purification of the recombinant protein.
Clone the SFRP1 gene into the expression vector and transform it with an appropriate expression host cell (e.g., E. coli);
Select the appropriate medium and conditions for large-scale culture, and purify the target protein using an affinity chromatography column (e.g., nickel column);
Samples were analyzed for purity and expression of the protein of interest using SDS-PAGE and Western blot.
Step 2: Functional analysis.
The cell lines or primary cells cultured in vitro were divided into experimental group and control group by cell experiment.
Experimental group: SFRP1 recombinant protein was added to the culture medium, and the expression changes of genes related to Wnt signaling pathway, such as -catenin, were observed.
Control group: SFRP1 recombinant protein was not added, and the expression changes of genes related to Wnt signaling pathway were observed.
The differences in gene or protein expression levels between the experimental group and the control group were analyzed by real-time PCR or Western blot.
4.Analysis of results:
The purified SFRP1 recombinant protein was obtained in step 1, and its purity and expression were verified by SDS-PAGE and Western blot.
In the cell culture experiment in step 2, it was observed that the genes related to the Wnt signaling pathway expressed in the experimental group changed compared with the control group, such as the decrease in the expression level of -catenin.
Further analysis showed that the SFRP1 recombinant protein inhibited the transmission of Wnt signaling by binding to WNT protein, which may be involved in the occurrence and development of WNT signaling, providing a new mechanism for the regulation of WNT signaling pathway.
Through the above experimental design and analysis, we were able to further study the regulatory role of SFRP1 recombinant protein in the Wnt signaling pathway and related mechanisms, providing important data and references for in-depth understanding of the pathogenesis mechanism.