Lamellar extracellular matrix (ECM) material enriched with vascular endothelial growth factor a (VEGFa) as well as preparation method and application thereof
A growth factor and vascular endothelial technology, applied in the field of extracellular matrix preparation, can solve the problems of insufficient application of treatment, limited quantity, high cost, etc.
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[0035] One aspect of the embodiments of the present invention provides a method for preparing a sheet-like extracellular matrix (hereinafter referred to as "ECM") material rich in vascular endothelial growth factor a (hereinafter referred to as "VEGFa"), which includes:
[0036] Constructing a recombinant vector containing a CBD-VEGFa fusion gene, and then using the recombinant vector to introduce the CBD-VEGFa fusion gene into a host cell, and then constructing a recombinant cell capable of expressing the vascular endothelial growth factor a;
[0037] Inoculating the recombinant cells on a culture dish for culturing;
[0038] The recombinant cells attached to the culture dish were freeze-dried to obtain a sheet-like extracellular matrix rich in vascular endothelial growth factor a.
[0039] In some preferred embodiments, the preparation method specifically includes: connecting the CBD-VEGFa fusion gene to the multiple cloning site of the pLVX-IRES-Puro vector, thereby constru...
Embodiment 1
[0056] The preparation method of a kind of extracellular matrix rich in vascular endothelial growth factor-a provided by this embodiment, the main technical process can be divided into four steps (see figure 1 Shown): 1. Vector construction; 2. Cell construction; 3. ECM preparation; 4. ECM performance testing.
[0057] Each step is described in detail below:
[0058] 1. Carrier Construction
[0059] (1) Select the appropriate carrier skeleton. The present invention selects the lentiviral vector pLVX-IRES-Puro (hereinafter referred to as PLVX) as the basic skeleton of the constructed recombinant vector.
[0060] (2) The National Center of Biotechnology Information (NCBI) website (https: / / www.ncbi.nlm.nih.gov / ) found the coding sequence (codingsequence, CDS) sequence of VEGFa and CBD.
[0061] (3) Use the Primer5 primer design software to design the primers required for VEGFa and CBD, and then submit them to the company to synthesize the primers. The list of primers is shown ...
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