Application of cell penetrating peptide Tat-mediated growth factor in transdermal transfer
A growth factor-mediated technology, applied in the field of transdermal drug delivery system and cosmetics, can solve the problems of difficult-to-explain macromolecular substances, and achieve the effect of increasing the transdermal rate and quantity
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Embodiment 1
[0021] Expression and purification of HIV-1-containing transactivator protein Tat protein transduction domain (Tat-protein transductiondomain, Tat-PTD 49-57 ) (Arg-Lys-Lys-Arg-Arg-Gln-Arg-Arg-Arg) and human epidermal growth factor fusion protein Tat-EGF. The gene fragment encoding Tat-EGF fusion protein was obtained by PCR method, and a 6×His tag was added at the 3' end. The amplified product was treated with restriction endonucleases BamHI and NdeI and connected to the Escherichia coli expression plasmid pET-3c (Novagen, co. Ltd., USA), positive clones were screened with ampicillin (final concentration 100ug / ml). The correct recombinant plasmid identified by sequencing was extracted and transformed into Escherichia coli BL21(DE3) (Novagen, co. Ltd., USA), and the transformants were selected by ampicillin resistance (final concentration 100ug / ml). At 37°C, after being induced by IPTG (final concentration: 1mmol / L) for 4 hours, the fusion protein was harvested and separated a...
Embodiment 2
[0023] Dissolve 0.1 g of hyaluronic acid in about 80 ml of double distilled water, then add heparin sodium (1 / 4 of the amount of Tat-EGF fusion protein), PEG4000.2 g, stir well and then autoclave (0.1Mpa, 121°C for 15 minutes ), cooled to room temperature. Add the Tat-EGF fusion protein with a purity of more than 98% obtained in Example 1 to the above-mentioned mixed solution after filtering and sterilizing through a 0.22 μm microporous membrane, adjust the pH to 6.8, and add sterilized double-distilled water to 100ml .
Embodiment 3
[0025] Take 0.1g of water-soluble chitosan and dissolve it in an appropriate amount of glacial acetic acid (about 1ml), add about 50ml of double-distilled water, adjust the pH to 6, then add 0.1g of EDTA-2Na, 0.2g of PEG4000, add methylparaben and Each 0.01g of ethylparaben (dissolved by heating with an appropriate amount of water first), stirred evenly, then autoclaved (0.1Mpa, 121°C for 15 minutes), and cooled to room temperature. Add the Tat-EGF fusion protein with a purity of more than 98% obtained in Example 1 to the above-mentioned mixed solution after filtering and sterilizing through a 0.22 μm microporous membrane, adjust the pH to 6.8, and add sterilized double-distilled water to 100ml . Dispensed in a spray bottle.
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