Genome Editing in Large Animals

Abstract

Genome editing in large animals has tremendous practical applications, from more accurate models for medical research through improved animal welfare and production efficiency. Although genetic modification in large animals has a 30 year history, until recently technical issues limited its utility. The original methods – pronuclear injection and integrating viruses – were plagued with problems associated with low efficiency, silencing, poor regulation of gene expression, and variability associated with random integration. With the advent of site specific nucleases such as TALEN and CRISPR/Cas9, precision editing became possible. When used on their own, these can be used to truncate or knockout genes through non-homologous end joining (NHEJ) with relatively high efficiency. When used with a template containing desired gene edits, these can be used to allow insertion of any desired changes to the genome through homologous recombination (HR) with substantially lower efficiency. Consideration must be given to the issues of marker sets and off-target effects. Somatic cell nuclear transfer is most commonly used to create animals from gene edited cells, but direct zygote injection and use of spermatogonial stem cells are alternatives under development. In developing gene editing projects, priority must be given to understanding the potential for off-target or unexpected effects of planned edits, which have been common in the past. Because of the increasing technical sophistication with which it can be accomplished, genome editing is poised to revolutionize large animal genetics, but attention must be paid to the underlying biology in order to maximize benefit.

The ethics of genome editing in non-human animals: a systematic review of reasons reported in the academic literature

“…Proactive and multidisciplinary collaboration can both advance these technological developments and the academic discourse about them, allowing us to go beyond rhetoric of promises or fears and positioning their ethical analysis in real-world practices [145,155]. CRISPR could help to meet the challenge of producing more food more sustainably to ensure the future global population can be fed 6 [34,78,79,[141][142][143] various (ZFN, TALEN); genetic modification could improve human health through the provision of new medicines and therapies 4 [26,126,133,140] various (TALEN, CRISPR) could enable genome engineering in non-human primates; this could be considered ethically problematic, but it is much more ethically problematic to watch people die who could be saved 1 [57] CRISPR could be used to create a chicken strain with low allergenicity 1 [126] CRISPR against (n ¼…”

“…could decrease animal suffering in dairy farming by creating dehorned cattle, preventing invasive and painful dehorning 10 [9,19,78 -80,85,96,126,139,140] various (ZFNs, TALEN, CRISPR) could counter welfare problems by creating the so-called diminished animals in which the ability to sense pain is impaired 8 [78,112,115,116,[119][120][121]137] genome editing (CRISPR); genetic engineering/modification could increase animal health and welfare by providing animals with disease resistance 8 [78,80,96,98,126,133,135,139] [19,137] various (ZFNs, TALEN, CRISPR); genetic engineering could be used to prevent the killing of day-old male chicks 2 [100,126] CRISPR; genetic modification re the possible creation of animals with welfare problems: if they have a life worth living we cannot say that they are worse off owing to the genetic modification, for if they had not been created with genetic modification, they would not have existed at all 2 [115,118] genetic modification re off-target effects: could result in fewer off-target effects than previous techniques, which could improve welfare of genetically modified animals 1 [9] CRISPR could reduce the numbers of animals used to create model organisms compared to traditional methods, which typically sacrifice many animals before achieving the desired genotype and phenotype 1 [110] CRISPR could remove known harmful recessive alleles that impair fertility or health and in that sense repair accumulated damage in the genome of breeding animals 1 [96] various (ZFNs, TALEN, CRISPR) could prevent wild animal suffering by using genome editing to change reproductive behaviour; the harm that would be prevented by doing so would outweigh the harm of developing and testing these strategies 1 [114] CRISPR could lead us to ignore the predicament of the animal and to accept negative effects on animal welfare for the sake of other goals; however, this concern may be addressed by using less drastic gene drive designs and using these to promote animal welfare 1…”

Translational Advances in the Field of Pulmonary Hypertension Molecular Medicine of Pulmonary Arterial Hypertension. From Population Genetics to Precision Medicine and Gene Editing

“…These have multiple potential applications in pulmonary hypertension research and therapy, from understanding the specific variants arising from "omics" studies to designing cells for targeted drug delivery applications, to the repair of mutations. Two relatively new technologies allow targeting of specific nucleotides: TALEN and CRISPR; each of these has multiple related technologies (51). TALEN, and the related technologies of zinc finger nucleases (52) Figure 2.…”

Non-Coding RNA Roles in Ruminant Mammary Gland Development and Lactation

“…Genome engineering has been considered as the next genomic revolution [144], and it is expected to significantly improve livestock production by precision genome editing [145][146][147] favouring markers associated with improved productivity, reproduction and health status. The history of genome editing in livestock has been extensively reviewed [145,[148][149][150].…”