A Method for Generating Transgenic Frog Embryos

Ishibashi, Shoko; Kroll, Kristen L.; Amaya, Enrique

doi:10.1007/978-1-60327-483-8_31

Cited by 45 publications

(16 citation statements)

References 44 publications

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“…The SV/Venus and SV/Venus/cSCLϩ19 constructs were generated by replacement of the LacZ reporter gene in SV/Lac and SV/Lac/ cSCLϩ19 by the Venus sequence from Venus/pCS2 (38) (a kind gift from Atsushi Miyawaki, Saitama, Japan). Double transgenic Xenopus laevis embryos were generated using restriction enzyme-mediated integration (27,31), using either SV/Venus or SV/Venus/cSCLϩ19 and ␥-crystallin-mCherry, which was derived from ␥-crystallin-green fluorescent protein (GFP) (6). The plasmids were linearized with SalI prior to the transgenesis protocol.…”

Section: Methodsmentioning

confidence: 99%

cis-Regulatory Remodeling of the SCL Locus during Vertebrate Evolution

Göttgens

Ferreira

Sánchez

et al. 2010

Molecular and Cellular Biology

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Development progresses through a sequence of cellular identities which are determined by the activities of networks of transcription factor genes. Alterations in cis-regulatory elements of these genes play a major role in evolutionary change, but little is known about the mechanisms responsible for maintaining conserved patterns of gene expression. We have studied the evolution of cis-regulatory mechanisms controlling the SCL gene, which encodes a key transcriptional regulator of blood, vasculature, and brain development and exhibits conserved function and pattern of expression throughout vertebrate evolution. SCL cis-regulatory elements are conserved between frog and chicken but accrued alterations at an accelerated rate between 310 and 200 million years ago, with subsequent fixation of a new cis-regulatory pattern at the beginning of the mammalian radiation. As a consequence, orthologous elements shared by mammals and lower vertebrates exhibit functional differences and binding site turnover between widely separated cis-regulatory modules. However, the net effect of these alterations is constancy of overall regulatory inputs and of expression pattern. Our data demonstrate remarkable cis-regulatory remodelling across the SCL locus and indicate that stable patterns of expression can mask extensive regulatory change. These insights illuminate our understanding of vertebrate evolution.Spatiotemporal changes in gene expression patterns represent one of the main driving forces of evolution (reviewed in references 10 and 41), with morphological divergence being linked to mutations of specific cis-regulatory elements in a growing number of examples (3,21,29,35,42). Conversely, evolutionarily conserved features are thought to reflect conserved gene expression patterns, but little is known about the mechanisms responsible for such stability. Most evidence comes from invertebrate studies and points to disparate mechanisms. For example, in the urochordate Ciona, orthologous muscle-specific enhancers are highly conserved across 270 million years (7), whereas orthologues of the Drosophila evenskipped stripe 2 enhancer exhibit multiple differences in transcription factor binding sites and yet have maintained the same spatiotemporal activity (33, 34). In vertebrates, the cis-regulatory elements of many regulatory genes are not conserved in comparisons between mammals and lower vertebrates (1, 14, 49), and little is known about the mechanisms responsible for maintaining stable gene expression patterns.The SCL gene encodes a basic helix-loop-helix (bHLH) transcription factor with pleiotropic and nonredundant roles in the development of the hematopoietic (40, 43), vascular (20, 36), and central nervous systems (5,20). Systematic dissection of cis-regulatory mechanisms operating at the murine SCL locus has identified multiple elements, each of which directs expression to a subdomain of the normal SCL expression pattern in transgenic mice. The two promoters (1a and 1b) have no hematopoietic activity (46), but three distal element...

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Section: Methodsmentioning

confidence: 99%

cis-Regulatory Remodeling of the SCL Locus during Vertebrate Evolution

Göttgens

Ferreira

Sánchez

et al. 2010

Molecular and Cellular Biology

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“…There are several ready-to-use practical manuals and reference books that describe various aspects of transgenic and knockout mouse technologies (2, 14, 45). Finally, species other than mouse have also been successfully used for transgenesis including chicken (46, 47), cow (48–50), fish (salmon and zebra fish) (51–54), frog (55, 56), goat (57–59), sheep (50, 60), pig (61, 62), rabbit (63–65), rat and monkey (67, 68). However, several methodological refinements are ongoing in several laboratories to routinely achieve transgenesis in species other than rodents.…”

Section: Methodsmentioning

confidence: 99%

Transgenic Mouse Technology: Principles and Methods

Kumar

Larson

Wang

et al. 2009

Methods in Molecular Biology

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Introduction of foreign DNA into the mouse germ line is considered a major technical advancement in the fields of developmental biology and genetics. This technology now referred to as transgenic mouse technology has revolutionized virtually all fields of biology and provided new genetic approaches to model many human diseases in a whole animal context. Several hundreds of transgenic lines with expression of foreign genes specifically targeted to desired organelles/cells/tissues have been characterized. Further, the ability to spatio-temporally inactivate or activate gene expression in vivo using the “Cre-lox” technology has recently emerged as a powerful approach to understand various developmental processes including those relevant to molecular endocrinology. In this chapter, we will discuss the principles of transgenic mouse technology, and describe detailed methodology standardized at our Institute.

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“…Many animal species have been genetically modified, including fish (Ozato et al, 1986), frogs (Ishibashi et al, 2008), rats (Agca et al, 2008), mice (Gordon and Ruddle, 1981;Joyner and Sedivy, 2000), cattle (Bondioli et al, 1991), birds (Etches and Verrinder Gibbins, 1997), and pigs (Kragh et al, 2009). The protocols to obtain such animals are diverse, but all consist of random integration of the exogenous gene into the genome of the animal.…”

Section: Types Of Genetically Modified Animals and How They Are Produmentioning

confidence: 99%

Review article Genetically modified animals for use in research and biotechnology

Chaible¹,

Corat²,

Abdelhay³

et al. 2010

Genet. Mol. Res.

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ABSTRACT. Transgenic animals are used extensively in the study of in vivo gene function, as models for human diseases and in the production of biopharmaceuticals. The technology behind obtaining these animals involves molecular biology techniques, cell culture and embryo manipulation; the mouse is the species most widely used as an experimental model. In scientific research, diverse models are available as tools for the elucidation of gene function, such as transgenic animals, knockout and conditional knockout animals, knock-in animals, humanized animals, and knockdown animals. We examined the evolution of the science for the development of these animals, as well as the techniques currently used in obtaining these animal models. We review the phenotypic techniques used for elucidation of alterations caused by genetic modification. We also investigated the role of genetically modified animals in the biotechnology industry, where they promise a revolution in obtaining heterologous proteins through natural secretions, such as milk, increas- ing the scale of production and facilitating purification, thereby lowering the cost of production of hormones, growth factors and enzymes.

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A Method for Generating Transgenic Frog Embryos

Cited by 45 publications

References 44 publications

cis-Regulatory Remodeling of the SCL Locus during Vertebrate Evolution

cis-Regulatory Remodeling of the SCL Locus during Vertebrate Evolution

Transgenic Mouse Technology: Principles and Methods

Review article Genetically modified animals for use in research and biotechnology

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