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Method for constructing zebrafish kidney progenitor cell marker transgenic line

A kind of kidney progenitor cell, construction method technology, applied in the field of construction of zebrafish kidney progenitor cell marker transgenic line

Active Publication Date: 2019-10-08
THE SECOND AFFILIATED HOSPITAL ARMY MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is currently no method that can efficiently label and facilitate gene overexpression and gene suppression in kidney progenitor cells

Method used

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  • Method for constructing zebrafish kidney progenitor cell marker transgenic line
  • Method for constructing zebrafish kidney progenitor cell marker transgenic line
  • Method for constructing zebrafish kidney progenitor cell marker transgenic line

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1 Six2a: Construction of GFF transgenic fish line

[0037] 1) six2 knock-in target screening:

[0038] a. Target selection and confirmation (on the last intron):

[0039] Through the principle of target design, 8 targets were found on the last intron of the six2 gene (see the gene nucleotide sequence SEQ ID NO.1) as follows Target1-Target8, respectively, the cutting efficiency was verified by genome sequencing, and the selected most efficient target.

[0040] Target design:

[0041]

[0042] b. Cutting efficiency verification:

[0043] After injecting 8 targets into zebrafish embryos respectively, the 3dpf juvenile genome was extracted and sequenced. Only target 6 and target 7 showed disordered peaks. figure 1 and figure 2 ;

[0044] c. Cloning verification cutting efficiency and cutting form:

[0045] Target 6: Among the 15 valid sequencing results, 2 had double peaks and were unusable, 12 had deletions or additions, and 1 had no mutation; therefore,...

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Abstract

The invention discloses a method for constructing a zebrafish kidney progenitor cell marker transgenic line. The method comprises the following steps: a gene knock-in method is used to construct a transgenic fish line of Six2a:GFF in a zebrafish, wherein the fish line hybridizes with UAS:GFP, UAS:Dendra2-NTR to specifically label kidney progenitor cells in the kidney or delete kidney progenitor cells, as well as overexpression and inhibition of the genes are carried out in kidney progenitor cells. The line can be used for high-throughput drug screening to provide the method to create kidney progenitors in human kidneys.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for constructing a marker transgenic line of zebrafish kidney progenitor cells. Background technique [0002] The strong regenerative capacity of the zebrafish kidney is due to the presence of renal progenitor cells in the zebrafish kidney. Research on zebrafish renal progenitor cells will provide new methods or drugs for the treatment of kidney diseases. However, there is currently no method for efficient labeling and convenient gene overexpression and gene suppression in renal progenitor cells. [0003] The inventors have successfully constructed a Six2a:GFF transgenic fish line in zebrafish by gene knock-in, which can specifically mark kidney progenitors in the kidney by crossing the fish line with UAS:GFP, UAS:Dendra2-NTR and other fish lines cells or delete renal progenitor cells, as well as overexpress and repress genes in renal progenitor cells. At the ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/85C12N15/90A01K67/027A61K49/00C12R1/91
CPCA01K67/0275A01K2217/072A01K2227/40A61K49/0008C12N15/85C12N15/902C12N2800/106
Inventor 刘赤喻婷傅瑶
Owner THE SECOND AFFILIATED HOSPITAL ARMY MEDICAL UNIV
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