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Expression method and application of ALB fusion protein

A technology of fusion protein and expression method, which is applied in the field of expression of ALB fusion protein to achieve high differentiation efficiency

Pending Publication Date: 2022-05-13
SHENZHEN SANQI BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, which expression method of ALB fusion protein can have a better effect, and obtain high-purity hepatocytes by sorting, and will not interfere with the metabolism of hepatocytes. There is no relevant literature report so far.

Method used

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  • Expression method and application of ALB fusion protein
  • Expression method and application of ALB fusion protein
  • Expression method and application of ALB fusion protein

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Embodiment Construction

[0054] The preferred embodiments of the present invention will be further described in detail below.

[0055] A method for expressing an ALB fusion protein, which adds one or more fusion proteins to the human ALB gene through one step. Such fusion proteins can be reporter genes (including but not limited to fluorescent protein genes, cell surface antigens, luciferase genes, chloramphenicol acetyltransferase genes, beta-galactosidase genes, secreted human placental alkaline phosphatase) genes, PET reporter genes, etc.), or functional genes. The fusion protein and ALB are connected by a 2A linking peptide, and the insertion position of the fusion protein is before the ALB stop codon. Two reporter genes were added to ALB through the 2A linking element before the ALB stop codon, but they did not interfere with the expression of endogenous ALB, and these two reporter genes were non-similar reporter genes, and no other screening genes were introduced (such as resistance gene or fl...

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Abstract

The invention provides an expression method and application of an ALB fusion protein, and the expression method comprises the following steps: cutting a genome by using nuclease, and simultaneously adding two fusion protein genes to an ALB gene through a gene co-expression connection element before a termination codon of the ALB gene of a human pluripotent stem cell or liver cell; the reporter gene is a fluorescent protein gene, a cell surface antigen, a luciferase gene, a chloramphenicol acetyltransferase gene, a beta-galactosidase gene, a secretory human placenta alkaline phosphatase gene or a PET reporter gene; human pluripotent stem cells are differentiated into liver cells through a directional differentiation technology. By adopting the technical scheme provided by the invention, the reporter gene or other functional genes can be efficiently connected with the ALB target gene, and the reporter gene or functional genes can be co-expressed with the target gene, so that specific reporter genes or functional proteins can be expressed in hepatocytes or hepatocyte-like cells.

Description

technical field [0001] The invention belongs to the technical field of molecular biology, and in particular relates to an expression method and application of an ALB fusion protein. Background technique [0002] Human induced pluripotent stem cells (iPSCs) can be differentiated into various cell types, including hepatocyte-like cells (iHeps). Since human iPSCs can expand indefinitely, they provide an unlimited number of liver cells for drug screening, liver toxicity testing, and regenerative medicine for cell replacement therapy. However, there is still a gap between the maturity of iHeps obtained from existing differentiation protocols and adult hepatocytes, and the cells in the differentiated iHeps population are heterogeneous and contain iHeps at different differentiation stages and other cell types. Cell lineage-specific fluorescent proteins or other reporter genes are particularly important for iPSCs. ALB is a gene encoding human albumin, which is specifically express...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/85C12N15/62C12N15/113C12N5/10C12N5/071
CPCC12N15/85C07K14/765C07K14/70517C07K14/70532C12N15/113C12N5/0696C12N5/067C12N2310/20C12N2510/00C12N2800/107C07K2319/60
Inventor 杨佳银付建江利香杨波
Owner SHENZHEN SANQI BIOTECH
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