Somatic Cell Nuclear Transfer in Zebrafish

Siripattarapravat, Kannika; Pinmee, Boonya; Cibelli, José B.

doi:10.1016/b978-0-12-386541-0.00016-3

Cited by 6 publications

(7 citation statements)

References 89 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In zebrafish, egg activation spontaneously occurs when the egg contacts hypotonic water (Wu and Kinsey, 2000) and we have adopted this activation in the cloning protocol. This activation event triggers the resumption of meiosis and cytoplasmic rearrangement (Siripattarapravat et al, 2009). A spontaneously activated egg precedes morphological changes and forms a blastodisc, which, although is morphologically indistinguishable from a one-cell zygote, undergoes no further division.…”

Section: Introductionmentioning

confidence: 99%

Use of soluble sperm extract to improve cloning efficiency in zebrafish

Prukudom¹,

Perez²,

Cibelli³

et al. 2019

Int. J. Dev. Biol.

Self Cite

View full text Add to dashboard Cite

During somatic cell nuclear transfer (SCNT), egg activation is required to initiate embryonic development. In zebrafish cloning, the reconstructed egg is activated by exposing it to hypotonic water. Egg activation using water-only is not capable of activating the same intracellular calcium release as fertilization which is required for proper embryonic development. Here we test whether the use of soluble sperm extract (SSE) can properly modulate the activation of reconstructed eggs during SCNT. We microinjected SSE from genomic-inactivated zebrafish sperm into unfertilized eggs and reconstructed eggs right after somatic cell nuclear transfer. We also evaluated the most effective approach for SSE microinjection. Microinjection of SSE (with 0.68 mg/ml of protein concentration) into non-activated eggs through the micropyle induced parthenogenetic development beyond the blastula stage, whereas all water-only activated eggs failed to enter the cleavage period. Microinjection of SSE at 1 mg/ml of protein concentration into non-activated reconstructed egg improved the developmental rate of cloned embryos in comparison to non-injected control clones. The cumulative survival time of cloned embryos injected with SSE was significantly longer than reconstructed eggs activated following sham injection (P<0.01). No significant difference was found among controls (P=0.32). SSE benefits both parthenogenesis and the survival cloned embryos which have never been reported in zebrafish. Further work is necessary to define the functional component(s) of SSE as well as the physiological pathway, to understand its principle of action and advance the utilization of SSE in cloning.

show abstract

Section: Introductionmentioning

confidence: 99%

Use of soluble sperm extract to improve cloning efficiency in zebrafish

Prukudom¹,

Perez²,

Cibelli³

et al. 2019

Int. J. Dev. Biol.

Self Cite

View full text Add to dashboard Cite

show abstract

“…There is an emerging awareness, however, thatholding these general cellular properties and technical aspects of the reprogramming regimen constant does little to hold results constant. This is observed in reprogramming experiments using either indirect means such as somatic cell nuclear transfer (SCNT -Campbell et al 1996;Cibelli et al, 1998;Siripattarapravat et al, 2009;Wakayama et al, 1998), or direct methods such as plasmid transduction or infection with recombinant retroviruses expressing transcription factors (Davis, Weintraub, and Lassar, 1987;Takahashi and Yamanaka, 2006;Vierbuchen et al, 2010;Ieda et al, 2010;Huang et al, 2011). One of the earliest studies of direct reprogramming, describing the conversion of mouse fibroblasts to skeletal muscle myotubes by transduction of the myogenic transcription factor MyoD1, found that reprogramming was not uniform across all cell lines.…”

Section: Introductionmentioning

confidence: 99%

Reprogramming to the nervous system: a computational and candidate gene approach

Alicea¹

2017

Preprint

View full text Add to dashboard Cite

The creation of stem-like cells, neuronal cells, and skeletal muscle fibers from a generic somatic precursor phenotype has many potential applications. These uses range from cell therapy to disease modeling. The enabling methodology for these applications is known as direct cellular reprogramming. While the biological underpinnings of cellular reprogramming go back to the work of Gurdon and other developmental biologists, the direct approach is a rather recent development.Therefore, our understanding of the reprogramming process is largely based on isolated findings and interesting results. A true synthesis, particularly from a systems perspective, is lacking. In this document, I will attempt to build toward an intellectual synthesis of direct reprogramming by critically examining four types of phenotypic conversion that result in production of nervous system components: induced pluripotency (iPS), induced neuronal (iN), induced skeletal muscle (iSM), and induced cardiomyocyte (iCM). Since potential applications range from tools for basic science to disease modeling and bionic technologies, the need for a common context is essential. This intellectual synthesis will be defined through several research endeavors.The first investigation introduces a set of experiments in which multiple fibroblast cell lines are converted to two terminal phenotypes: iN and iSM. The efficiency and infectability of cells subjected to each reprogramming regimen are then compared both statistically and quantitatively. This set of experiments also resulted in the development of novel analytical methods for measuring reprogramming efficiency and infectability.The second investigation features a critical review and statistical analysis of iPS reprogramming, specifically when compared to indirect reprogramming (SCNT-ES) and related stem-like cells. The third investigation is a review and theoretical synthesis which stakes out new directions in our understanding of the direct reprogramming process, including recent computational modeling endeavors and results from the iPS, iN and induced cardiomyocyte (iCM) experiments.To further unify the outcomes of these studies, additional results related to Chapter 2 and directions for future research will be presented. The additional results will allow for further interpretation and insight into the role of diversity in direct reprogramming. These future directions include both experimental approaches (a technique called mechanism disruption) and computational approaches (preliminary results for an agent-based population-level approximation of direct reprogramming). The insights provided here will hopefully provide a framework for theoretical development and a guide for traditional biologists and systems biologists alike.Several components of this volume have been previously published. Chapter 2 is published as an article in Stem Cells and Development (Volume 22, Issue 19). Chapter 3 is published as a book chapter (Chapter 37) in the book “Principles in Cloning, Volume III” (Elsevier). Special thanks go to Dr. Steven Suhr and Dr. Jose Cibelli, who are co-authors for Chapter 2 and Chapter 3, respectively.

show abstract

“…There is an emerging awareness, however, that even when these general cellular properties and technical aspects of the reprogramming regimen are held constant, variation may still be the rule rather than the exception. This is observed in reprogramming experiments using either indirect means such as somatic cell nuclear transfer [1][2][3][4], or direct methods such as plasmid transduction or infection with recombinant retroviruses expressing transcription factors [5][6][7][8][9]. One of the earliest studies of direct reprogramming, describing the conversion of mouse fibroblasts to skeletal muscle myotubes by transduction of the myogenic transcription factor MyoD1, found that reprogramming was not uniform across all cell lines.…”

Section: Introductionmentioning

confidence: 99%

Defining the Diversity of Phenotypic Respecification Using Multiple Cell Lines and Reprogramming Regimens

Alicea

Murthy

Keaton

et al. 2013

Stem Cells and Development

View full text Add to dashboard Cite

To better understand the basis of variation in cellular reprogramming, we performed experiments with two primary objectives: first, to determine the degree of difference, if any, in reprogramming efficiency among cells lines of a similar type after accounting for technical variables, and second, to compare the efficiency of conversion of multiple similar cell lines to two separate reprogramming regimens-induced neurons and induced skeletal muscle. Using two reprogramming regimens, it could be determined whether converted cells are likely derived from a distinct subpopulation that is generally susceptible to reprogramming or are derived from cells with an independent capacity for respecification to a given phenotype. Our results indicated that when technical components of the reprogramming regimen were accounted for, reprogramming efficiency was reproducible within a given primary fibroblast line but varied dramatically between lines. The disparity in reprogramming efficiency between lines was of sufficient magnitude to account for some discrepancies in published results. We also found that the efficiency of conversion to one phenotype was not predictive of reprogramming to the alternate phenotype, suggesting that the capacity for reprogramming does not arise from a specific subpopulation with a generally ''weak grip'' on cellular identity. Our findings suggest that parallel testing of multiple cell lines from several sources may be needed to accurately assess the efficiency of direct reprogramming procedures, and that testing a larger number of fibroblast lines-even lines with similar origins-is likely the most direct means of improving reprogramming efficiency.

show abstract

Somatic Cell Nuclear Transfer in Zebrafish

Cited by 6 publications

References 89 publications

Use of soluble sperm extract to improve cloning efficiency in zebrafish

Use of soluble sperm extract to improve cloning efficiency in zebrafish

Reprogramming to the nervous system: a computational and candidate gene approach

Defining the Diversity of Phenotypic Respecification Using Multiple Cell Lines and Reprogramming Regimens

Contact Info

Product

Resources

About