ZFNs animal model

NegotiableUpdate on 03/04

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Nature of the Manufacturer: Producers

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Overview

ZFNs animal model: Zinc finger nucleases (ZFNs) animal model is an experimental animal model constructed using zinc finger nucleases gene editing technology. Zinc finger nucleases are formed by the fusion of zinc finger proteins (ZFP) and nucleases (FokI). Zinc finger proteins can specifically bind to target DNA sequences, while nucleases are responsible for cutting DNA, introducing double strand breaks at specific gene loci, inducing cell repair mechanisms, and achieving gene knockout, knock in, or mutation.

Product Details

1ZFNs animal modelTechnical principles and core mechanisms

ZFNs areThe first generation of programmable gene editing toolsThrough integrationDNA binding domain(Zinc finger protein) andDNA cleavage domain(FokI nuclease) achieves targeted editing:

Zinc finger protein domain：

from30-34 amino acid repeat unitsComposition, each unit passes through12th and 13th variable residues (RVDs)Identify specific bases:

RVD type	Identify bases	specificity
NI	A	high
NG	T	high
HD	C	high
NN	G/A	moderate

Single Zinc Finger Identification3 bpConnect 3-6 targetable devices in series9-18 bpSequence.

FokI cutting domain：

needdimerizationTo activate the cutting function, ZFNs need to be designed in pairs (left arm/right arm), with the cutting site located between the binding sites of the two arms (5-7 bp interval).

Editing mechanism：

Double strand break (DSB) → cell initiated repair pathway:

Non homologous terminal junction (NHEJ)Random insertion/deletion (Indels) leads to gene knockout.
Homologous Directional Restoration (HDR)External repair templates (such as ssODN) are required to achieve point mutations or gene insertions.

technological breakthroughShou times implementationMammalian genome targeted editingIn 2005, the era of precise gene editing began.

IIZFNs animal modelProcess and Technology Optimization

（1） Standardized operating procedures

（2） Optimization of key technical parameters

step	Core Operations	Innovation optimization
Target design	Avoid high repetition/methylation areas, with a spacing of 5-7 bp	Software prediction (ZiFiT) improves integration efficiency
Zinc finger assembly	Modular series method：

3-6 zinc finger units in series
Golden Gate Clone (BsAI/BsmBI) | Built in 6 days with a success rate>80%|
|expression vector|Dual promoter vector (CMV/T7), supporting mRNA synthesis and protein expression | adaptable to multiple species (zebrafish, mice, pigs)|
|Delivery method|Microinjection of ZFNs mRNA into fertilized eggs
Somatic cell transfection+nuclear transfer (large animals) | Reduced off target mRNA delivery|
|Repair and Enhancement|Combined ssODN template+NHEJ inhibitor (SCR7) | HDR efficiency increased by 3 times|

3、 Multi species application cases and model advantages

（1） Typical Species and Disease Models

species	target gene	Disease Model	Efficiency/phenotype	research value
zebrafish	golden	albinism	95% F0 generation pigment deficiency	Functional screening of developmental genes
mouse	Rag2/IL2rg	Immunodeficiency model	Dual gene knockout, humanized transplantation platform	Research on tumor immunotherapy
pig	GGTA1	Xenomorph Guan transplantation model	Knocking out alpha Gal antigen to reduce hyperacute rejection	Humanized organ development
fruit fly	yellow	Body color mutation	Reproductive system mutation rate 5.7% (animal model)	Gene functional conservation verification

（2） Irreplaceable application scenarios

High GC region editing：

Without PAM sequence restrictions, it can target regions that CRISPR/Cas9 cannot edit.

Low off target rate requirement：

The off target rate in therapeutic editing is less than 0.1% (1-10% for CRISPR).

Large animal models：

The immunogenicity of pigs, cows and other species is lower than CRISPR.