One-step base editing in multiple genes by direct embryo injection for pig trait improvement

Song, Ruigao; Wang, Yu; Zheng, Qiantao; Cao, Chunwei; Wang, Yanfang; Zhao, Jianguo

doi:10.1007/s11427-021-2013-8

Cited by 22 publications

(9 citation statements)

References 40 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In addition, 80.0% of the single clones simultaneously contained desired stop codons in all these three genes (Table 1). In a recent report [35], hA3A-BE3-Y130F exhibited around 10-60% C−to−T editing efficiencies in PFFs for a three-gene simultaneous-editing experiment, and showed 55.6-100% editing efficiencies for single and multiple genes in porcine embryos. In our study, this CBE displayed 87.2-98.6% desired C−to−T mutation rates in each of the three tumor-suppressor genes.…”

Section: Discussionmentioning

confidence: 90%

Efficient Simultaneous Introduction of Premature Stop Codons in Three Tumor Suppressor Genes in PFFs via a Cytosine Base Editor

Jiang

Jing

Yang

et al. 2022

Genes

View full text Add to dashboard Cite

Base editing is an efficient and precise gene-editing technique, by which a single base can be changed without introducing double-strand breaks, and it is currently widely used in studies of various species. In this study, we used hA3A-BE3-Y130F to simultaneously introduce premature stop codons (TAG, TGA, and TAA) into three tumor suppressor genes, TP53, PTEN, and APC, in large white porcine fetal fibroblasts (PFFs). Among the isolated 290 single-cell colonies, 232 (80%) had premature stop codons in all the three genes. C−to−T conversion was found in 98.6%, 92.8%, and 87.2% of these cell colonies for TP53, PTEN, and APC, respectively. High frequencies of bystander C−to−T edits were observed within the editing window (positions 3–8), and there were nine (3.01%) clones with the designed simultaneous three-gene C−to−T conversion without bystander conversion. C−to−T conversion outside the editing window was found in 9.0%, 14.1%, and 26.2% of the 290 cell colonies for TP53, PTEN, and APC, respectively. Low-frequency C−to−G or C−to−A transversion occurred in APC. The mRNA levels of the three genes showed significant declines in triple-gene-mutant (Tri-Mut) cells as expected. No PTEN and a significantly lower (p < 0.05) APC protein expression were detected in Tri-Mut cells. Interestingly, the premature stop codon introduced into the TP53 gene did not eliminate the expression of its full-length protein in the Tri-Mut cells, suggesting that stop codon read-through occurred. Tri-Mut cells showed a significantly higher (p < 0.05) proliferation rate than WT cells. Furthermore, we identified 1418 differentially expressed genes (DEGs) between the Tri-Mut and WT groups, which were mainly involved in functions such as tumor progression, cell cycle, and DNA repair. This study indicates that hA3A-BE3-Y130F can be a powerful tool to create diverse knockout cell models without double-strand breaks (DSBs), with further possibilities to produce porcine models with various purposes.

show abstract

Section: Discussionmentioning

confidence: 90%

Efficient Simultaneous Introduction of Premature Stop Codons in Three Tumor Suppressor Genes in PFFs via a Cytosine Base Editor

Jiang

Jing

Yang

et al. 2022

Genes

View full text Add to dashboard Cite

show abstract

“…Additionally, different CRISPR-based strategies were applied to target the porcine MSTN gene at the in vitro level ,, and to produce live MSTN -edited pigs (Table ). The cytosine base editor (CBE) was also applied to edit the porcine MSTN gene and generate pigs harboring multiple point mutations (stop codons in CD163 and MSTN , and a beneficial allele in IGF2 ) . Of note, another strategy based on the homology-mediated end joining (HMEJ)-induced site-specific integration of an MSTN inhibitor, follistatin ( FST ), into the last codon of the porcine MSTN gene, was also applied to investigate its role in the increase of skeletal muscle mass .…”

Section: Targeting the Mstn Gene In Pigsmentioning

confidence: 99%

“…The cytosine base editor (CBE) was also applied to edit the porcine MSTN gene 92 and generate pigs harboring multiple point mutations (stop codons in CD163 and MSTN, and a beneficial allele in IGF2). 93 Of note, another strategy based on the homologymediated end joining (HMEJ)-induced site-specific integration of an MSTN inhibitor, follistatin (FST), into the last codon of the porcine MSTN gene, was also applied to investigate its role in the increase of skeletal muscle mass. 94 These studies demonstrate the generation of the double-muscling phenotype and improved meat production in MSTN-edited pigs.…”

Section: ■ Targeting the Mstn Gene In Pigsmentioning

confidence: 99%

When Less Is More: Targeting the Myostatin Gene in Livestock for Augmenting Meat Production

Kalds

Zhou

Huang

et al. 2023

J. Agric. Food Chem.

View full text Add to dashboard Cite

How to increase meat production is one of the main questions in animal breeding. Selection for improved body weight has been made and, due to recent genomic advances, naturally occurring variants that are responsible for controlling economically relevant phenotypes have been revealed. The myostatin (MSTN) gene, a superstar gene in animal breeding, was discovered as a negative controller of muscle mass. In some livestock species, natural mutations in the MSTN gene could generate the agriculturally desirable double-muscling phenotype. However, some other livestock species or breeds lack these desirable variants. Genetic modification, particularly gene editing, offers an unprecedented opportunity to induce or mimic naturally occurring mutations in livestock genomes. To date, various MSTN-edited livestock species have been generated using different gene modification tools. These MSTN gene-edited models have higher growth rates and increased muscle mass, suggesting the high potential of utilizing MSTN gene editing in animal breeding. Additionally, post-editing investigations in most livestock species support the favorable influence of targeting the MSTN gene on meat quantity and quality. In this Review, we provide a collective discussion on targeting the MSTN gene in livestock to further encourage its utilization opportunities. It is expected that, shortly, MSTN gene-edited livestock will be commercialized, and MSTN-edited meat will be on the tables of ordinary customers.

show abstract

“…The CBE system was first used to generate base editing pigs to establish larger animal models [ 70 ]. Subsequently, many groups have used CBE and ABE to knock out different porcine genes, such as GGTA, MSTN, CD163, GHR, and IGF2, for pig trait improvement [ 71 , 72 , 73 , 74 , 75 ]. BE3 and ABEmax generated goats and sheep with Socs2, GFG5, and BMPRIB mutations in Northwest A&F University [ 76 , 77 , 78 ].…”

Section: Gene Editing Tools and Their Application In Farm Animalsmentioning

confidence: 99%

Application of Gene Editing Technology in Resistance Breeding of Livestock

Wang

Huang

et al. 2022

Life

View full text Add to dashboard Cite

As a new genetic engineering technology, gene editing can precisely modify the specific gene sequence of the organism’s genome. In the last 10 years, with the rapid development of gene editing technology, zinc-finger nucleases (ZFNs), transcription activator-like endonucleases (TALENs), and CRISPR/Cas9 systems have been applied to modify endogenous genes in organisms accurately. Now, gene editing technology has been used in mice, zebrafish, pigs, cattle, goats, sheep, rabbits, monkeys, and other species. Breeding for disease-resistance in agricultural animals tends to be a difficult task for traditional breeding, but gene editing technology has made this easier. In this work, we overview the development and application of gene editing technology in the resistance breeding of livestock. Also, we further discuss the prospects and outlooks of gene editing technology in disease-resistance breeding.

show abstract

One-step base editing in multiple genes by direct embryo injection for pig trait improvement

Cited by 22 publications

References 40 publications

Efficient Simultaneous Introduction of Premature Stop Codons in Three Tumor Suppressor Genes in PFFs via a Cytosine Base Editor

Efficient Simultaneous Introduction of Premature Stop Codons in Three Tumor Suppressor Genes in PFFs via a Cytosine Base Editor

When Less Is More: Targeting the Myostatin Gene in Livestock for Augmenting Meat Production

Application of Gene Editing Technology in Resistance Breeding of Livestock

Contact Info

Product

Resources

About