A kind of polypeptide for preparing hydrogel
A technology for hydrogels and products, applied in the field of polypeptide screening, can solve problems such as mechanical property limitations, and achieve the effect of large application potential
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Embodiment 1
[0022] Example 1: Screening of polypeptides
[0023] 1.1: Hl-TSR-like omics information
[0024] Based on the comparison of the transcriptome databases of the foot disc and cylinder of the inventor's research group, it was found that the Hl-TSR-like gene fragment was a significantly up-regulated gene in the foot disc, and it was compared with the local sequence. It is classified as a protein of the thrombin-sensitizing protein-1 type 1 repeat (TSR) class.
[0025] The expression level (FPKM) of Hl-TSR-like in the anemone cylinder was 50.51, and the expression level (FPKM) in the foot disc was 11497.97, which belonged to genes differentially expressed in the sea anemone foot disc. The mass spectrometry data of Adnectin showed that 6 peptides were identified for Hl-TSR-like, four of which were unique to Hl-TSR-like.
[0026] 1.2: Hl-TSR-like RACE (Rapid-amplification of cDNA ends) technology
[0027] In RNA-Seq, the H1-TSR-like sequence was obtained due to gene splicing. In o...
Embodiment 2
[0050] Example 2: Construction of H1-TSR-like-pET28a recombinant expression system
[0051] Recombinant expression was carried out in Escherichia coli by constructing the expression system Hl-TSR-like-pET28a containing a hexahistidine tag. After adding the stop codon TAA at the 3' end, the forward primer and reverse primer were designed from the 5' end and 3' end, respectively. The forward primer contains a restriction endonuclease BamH I restriction site (GGATCC), and a protective base CG is added upstream of the BamH I restriction endonuclease site, thereby increasing the efficiency of restriction endonuclease digestion. Similarly, the reverse primer contains a restriction endonuclease Xho I cutting site (CTCGAG) and an expression guard base CCG is added upstream. The primer information required for amplification is as follows:
[0052] Upstream primer: 5'-CGGGATCCTGTCCAATTCATGGAGGATACG-3'
[0053] Downstream primer: 5'-CCGCTCGAGTTAGTAGGCTGGTGGAGGAGGTG-3'
[0054] The pu...
Embodiment 3
[0065] Embodiment 3: Characterization of the properties of H1-TSR-like polypeptide
[0066] 3.1: Preparation of hydrogel
[0067] Adjust the concentration of Hl-TSR-like protein after refolding in pH 8.5 and 20mM Tris-HCl to higher than 25mg / ml, transfer the concentrated protein solution to the mold, and let it stand at 16°C for 72h, the protein can self-polymerize into a hydrogel, and the resulting hydrogel has a shape such as Figure 5 shown.
[0068] The shape of the protein hydrogel is controllable, and the gel conditions in the molds of different materials remain unchanged, and samples of various shapes can be manufactured according to requirements. Another notable feature of hydrogel is its strong self-polymerization ability, such as Figure 5 As shown in (a), the hydrogel can be restored to its original shape within 5 min after being cut into two pieces, which is attributed to the strong interaction force between the protein interiors.
[0069] 3.2: Hydrogel Surface...
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