Method for transducing buffalo embryo with PEP-1 peptide concatemer mediated green fluorescent protein

Abstract

The invention relates to the technical field of targeting transport of cell penetrating peptides, in particular to a method for transducing a buffalo embryo with PEP-1 peptide concatemer mediated green fluorescent protein. The method comprises steps as follows: design and optimization of PEP-1-PEP-1-EGFP, construction of a prokaryotic expression vector pCZN1-PEP-1-PEP-1-EGFP, prokaryotic expression of the protein, protein transfection and the like. By means of genetic engineering, the PEP-1 concatemer mediated expression vector plasmid pCZN1-PEP-1-PEP-1-EGFP is constructed, the optimal condition for transducing buffalo embryo cells with the PEP-1 peptide concatemer mediated green fluorescent protein is explored, the green fluorescent protein can be transduced into the buffalo embryo accurately and efficiently, a research foundation is laid for study of target transduction of nutrient substances and target effects of drugs, and the mediation effect of PEP-1 concatemers can be exerted efficiently.

Description

[0001] 【Technical Field】 [0002] The present invention relates to the technical field of targeted transport of cell-penetrating peptides, in particular to a method for transducing green fluorescent protein into buffalo embryos by PEP-1 peptide concatemers. [0003] 【Background technique】 [0004] During the in vitro maturation and culture process, buffalo embryos have undergone the stages of fertilized egg, 2-cell, 4-cell, 8-cell, mulberry, blastocyst, and hatching blastocyst. In the early developmental stage of the buffalo embryo, the zona pellucida on the periphery is important for maintaining the embryo. The morphology and protection of the internal environment of the embryo play important functions, but at the same time, in the process of in vitro embryo production, the zona pellucida also hinders the embryo's uptake of artificially added biomolecules. Cell penetrating peptides (CPPs), generally no more than 30 amino acids in length, have the characteristics of carrying macromol...

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Problems solved by technology

For example, on pages 798-801 of Volume 27, Issue 7 of "Basic Medicine and Clinic" in July 2007, an article entitled "Cell Penetrating Peptide PEP-1 Mediates Enhanced Green Fluorescent Protein Transcending in Mice Membrane Transduction" paper, introduced the use of PEP-1 to mediate the transmembrane transduction of green fluorescent protein in mice. The concatemer was connected to green fluorescent protein, then connected to pET15b prokaryotic expression vector, injected into mice, and detected the area of ​​green fluorescent protein expression, proving that the PEP-1 carrier can transfer nutrients to the brain, heart muscle, liver, and spleen and kidney tissue, to prove that the PEP-1 carrier can transmembranely transduce nutrients to the vital organs of mice, but there is currently no report on the delivery of PEP-1 to buffalo embryos in vitro, delivery of nutritional factors, etc. There is no report on the use of PEP-1 tandem to deliver nutrient factors to buffalo embryos in vitro

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