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Methods in Cell Biology 2019Methods for microinjection into sea urchin eggs have become relatively easier because of the technical improvements by a number of researchers in the past decades.... (Review)
Review
Methods for microinjection into sea urchin eggs have become relatively easier because of the technical improvements by a number of researchers in the past decades. However, the size and the characteristics, such as the elasticity and toughness, of the eggs and embryos differ in species, so that we still need to modify the details of methods to adapt to each target. In this section, I list microinjection methods for three species: Hemicentrotus pulcherrimus, which has relatively tough eggs, Temnopleurus reevesii, which has slightly weak eggs, and Strongylocentrotus purpuratus, which is the most used species in sea urchin biology. In addition, I describe the methods for co-injection of morpholino anti-sense oligonucleotides and mRNAs, as well as the method for microinjection into blastomeres.
Topics: Animals; Blastomeres; Embryo, Nonmammalian; Microinjections; Oocytes; Sea Urchins
PubMed: 30777175
DOI: 10.1016/bs.mcb.2018.09.013 -
Cloning and Stem Cells 2001Last year marked the 20th anniversary of the invention of the term "transgenic" and the development of pronuclear microinjection, a straightforward technique designed to... (Review)
Review
Last year marked the 20th anniversary of the invention of the term "transgenic" and the development of pronuclear microinjection, a straightforward technique designed to transfer genetic information from nearly any living organism to mammals. After two decades of use, pronuclear microinjection protocols have changed little from the reliable, if not efficient, method described by Gordon and Ruddle. Experience has taught us that once microinjection skills are perfected there are only a few parameters one needs to be concerned about to successfully produce transgenic animals. Those parameters will be discussed, as will some new innovations that promise to finally increase efficiency of pronuclear microinjection methodology.
Topics: Animals; Animals, Genetically Modified; Cell Nucleus; Gene Transfer Techniques; Mammals; Microinjections; Nuclear Transfer Techniques; Oocytes
PubMed: 11945231
DOI: 10.1089/15362300152725936 -
Methods in Molecular Biology (Clifton,... 1999
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Methods in Molecular Biology (Clifton,... 2019Microinjection is a powerful tool for studying embryonic development and analyzing gene functions in fish. This technique was first applied to model species of fish such...
Microinjection is a powerful tool for studying embryonic development and analyzing gene functions in fish. This technique was first applied to model species of fish such as zebrafish and medaka whose egg chorions could be removed or softened before microinjection. Recent progress in genome editing using TALEN and CRISPR has opened the opportunity to analyze gene functions in a much wider range of fish including those important to marine aquaculture. Therefore, application of the microinjection technique is also required in these species. However, the characteristics of fish eggs vary widely between species and several technical difficulties need to be overcome in order to use microinjection in a wider range of species. To obtain consistent results with microinjection, an optimal method has to be developed for each target species. In this chapter, we describe the physical characteristics of the eggs of fish species that have been used in microinjection experiments in our laboratory and detail the microinjection system we developed for fish eggs with a hard chorion, such as those of marine species.
Topics: Animals; Animals, Genetically Modified; Embryonic Development; Female; Fishes; Gene Editing; Microinjections; Ovum
PubMed: 30353531
DOI: 10.1007/978-1-4939-8831-0_27 -
Current Opinion in Biotechnology Oct 2008Microinjection to single cells has been widely used in the studies of transduction-challenged cells, transgenic animal production, and in vitro fertilization to... (Review)
Review
Microinjection to single cells has been widely used in the studies of transduction-challenged cells, transgenic animal production, and in vitro fertilization to mechanically transfer DNAs, RNA interferences, sperms, proteins, peptides, and drugs. The advantages of microinjection include the precision of delivery dosage and timing, high efficiency of transduction as well as low cytotoxicity. However, manual microinjection is labor intensive and time consuming, which limits the application of this technique to large number of cells in a sample. New cell culture matrix ensuring all cells grow in a desired position and orientation is needed for application of high throughput automatic injection systems, which will significantly increase injection speed, cell survival, and success rates.
Topics: Drug Delivery Systems; Gene Transfer Techniques; Mechanical Phenomena; Microinjections
PubMed: 18725294
DOI: 10.1016/j.copbio.2008.07.005 -
Methods in Molecular Biology (Clifton,... 1999
Review
Topics: Animals; Microinjections; Oocytes; Xenopus laevis
PubMed: 10503229
DOI: 10.1385/1-59259-678-9:111 -
Methods in Molecular Biology (Clifton,... 2019Microinjection/micromanipulation is more than 100 years old. It is a technique that is instrumental in biomedical research and healthcare. Its longevity lies in its...
Microinjection/micromanipulation is more than 100 years old. It is a technique that is instrumental in biomedical research and healthcare. Its longevity lies in its preciseness in mechanical retrieval, or delivery of biological materials, which in some cases is simply necessary or more effective than other retrieval/delivery means. Microinjection is favored for its straightforwardness in transferring contents from micromolecules to macromolecules and from organelles to cells. Microinjection/micromanipulation has been practiced over the century like an art form. Variations in handlings and instruments can be tolerated to a surprising degree with satisfactory outcomes. Throughout the century, microinjection developed as an indispensable tool along with the evolution of biomedical fields: from transgenics to gene targeting, from animal cloning to human infertility treatment, from nuclease-guided genetic engineering to RNA-guided genome editing (Fig. 1). The birth of the CRISPRology rejuvenated microinjection. For microinjection/micromanipulation, the second century has already begun with the early arrival of computerized instrumentation and lately of the high-throughput nanomanipulators potentially operable by artificial intelligence. As we yin-yang both systemic and precision approaches in research and medicine, microinjection will no doubt continue to find its unique place in the future.
Topics: Animals; CRISPR-Cas Systems; Gene Editing; Gene Targeting; Genetic Engineering; History, 20th Century; History, 21st Century; Humans; Microinjections; Micromanipulation; Nanotechnology
PubMed: 30353505
DOI: 10.1007/978-1-4939-8831-0_1 -
Methods in Molecular Biology (Clifton,... 2020The ability to delete the function of an endogenous gene in the mouse was made possible by the development of embryonic stem (ES) cells, pluripotent cells that retain...
The ability to delete the function of an endogenous gene in the mouse was made possible by the development of embryonic stem (ES) cells, pluripotent cells that retain the ability to develop into all tissues of a developing embryo. The ability to genetically modify these cells followed, allowing targeted mutation of ES cells in vitro and the deletion of specific gene function. However, regardless of the simplicity or complexity of the genetic modification, all ES cells require injection into host embryos to establish pregnancies and result in chimeric mice. Blastocysts are commonly used as the host embryos for this purpose, as it is relatively easy to inject cells into the blastocoel cavity of the developing embryo. This chapter describes the procedure for injection of ES cells into blastocyst stage embryos for the generation of knockout mice.
Topics: Animals; Blastocyst; Chimera; Embryo Transfer; Embryonic Stem Cells; Female; Mice; Mice, Knockout; Microinjections; Pregnancy
PubMed: 31512208
DOI: 10.1007/978-1-4939-9837-1_6 -
BioEssays : News and Reviews in... Jun 2008Single-cell microinjection has been successfully used to deliver exogenous proteins, cDNA constructs, peptides, drugs and particles into transfection-challenged cells.... (Review)
Review
Single-cell microinjection has been successfully used to deliver exogenous proteins, cDNA constructs, peptides, drugs and particles into transfection-challenged cells. With precisely controlled delivery dosage and timing, microinjection has been used in many studies of primary cultured cells, transgenic animal production, in vitro fertilization and RNA inference. This review discusses the advantages and limits of microinjection as a mechanical delivery method and its applications to attached and suspended cells.
Topics: Animals; Cell Adhesion; Cells, Cultured; Cytological Techniques; Fluorescent Dyes; Genetic Techniques; Humans; Microinjections; Neurons; Transduction, Genetic
PubMed: 18478541
DOI: 10.1002/bies.20759 -
Methods in Cell Biology 2000
Review
Topics: Animals; Gene Transfer Techniques; Microinjections; Tetrahymena thermophila; Transformation, Genetic
PubMed: 10503212
DOI: 10.1016/s0091-679x(08)61551-4