The effect of pronuclear microinjection of DNA and culture in excised mouse oviducts on the development of porcine zygotes was assessed in this study. Precocious ovulation was induced in prepubertal gilts with pregnant mare's serum gonadotrophin and hCG. Zygotes received either pronuclear microinjection of buffer alone, buffer containing a DNA construct, or no microinjection. Zygotes were cultured in vitro in either modified Krebs-Ringer bicarbonate medium (KRB) for 144 h or in mouse oviduct (MO) explant culture with KRB for 48, 72, 96, or 120 h. Pronuclear microinjection of DNA resulted in a lower (P less than .05) cleavage index (CI) than did buffer or no microinjection (CI 2.16 +/- .10 vs 2.80 +/- .13 and 2.93 +/- .10). The CI loss was greatest for DNA-injected zygotes at the two-cell stage of development. Coculture of zygotes in MO resulted in a higher CI (P less than .01) than did culture in KRB. Culture in MO for 72 h was the most beneficial system compared with MO for 48, 96, or 120 h (P less than .05; CI 3.25 +/- .12 vs 2.66 +/- .18, 2.79 +/- .14, and 2.40 +/- .14, respectively). Microinjection of DNA, not merely the mechanical procedure, was detrimental to early zygote development and may be the cause of low pregnancy rates.
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Bio Protoc
December 2024
Division of Evolutionary Developmental Biology, National Institute for Basic Biology, Okazaki, Japan.
In this paper, we present a detailed protocol for microinjecting DNA, RNA, or protein solutions into fertilized eggs of the multicolored Asian ladybird beetle, Harmonia axyridis, under a stereomicroscope equipped with an injection apparatus. is an emerging model organism for studying various biological fields, showing intraspecific polymorphisms exhibiting highly diverse color patterns on the elytra. Here, we describe how to rear ladybird beetles in a laboratory and obtain fertilized eggs for microinjection experiments.
View Article and Find Full Text PDFAnimal Model Exp Med
December 2024
Frontiers Science Center for Molecular Design Breeding (MOE), State Key Laboratory of Animal Biotech Breeding, College of Biological Sciences, China Agricultural University, Beijing, China.
Background: Traditional DNA microinjection methods used in mammals are difficult to apply to avian species due to their unique reproductive characteristics. Genetic manipulation in chickens, particularly involving immature follicles within living ovaries, has not been extensively explored. This study seeks to establish an efficient method for generating transgenic chickens through ovarian injection, potentially bypassing the challenges associated with primordial germ cell (PGC) manipulation and fertilized egg microinjection.
View Article and Find Full Text PDFPeptides
January 2025
Department of Special Medicine, School of Basic Medicine, Qingdao University, Qingdao 266000, China. Electronic address:
Biochim Biophys Acta Mol Cell Res
February 2025
Friedrich-Alexander-Universität Erlangen-Nürnberg, Department of Anesthesiology, Krankenhausstraße 12, 91054 Erlangen, Germany. Electronic address:
Objective: Transgenic mice with fluorescent protein (FP) reporters take full advantage of new in vivo imaging technologies. Therefore, we generated a TRPC5- and a TRPA1-reporter mouse based on FP C-terminal fusion, providing us with better alternatives for studying the physiology, interaction and coeffectors of these two TRP channels at the cellular and tissue level.
Methods: We generated transgenic constructs of the murine TRPC5- and TRPA1-gene with a 3*GGGGS linker and C-terminal fusion to mCherry and mTagBFP, respectively.
Biomolecules
November 2024
Experimental Mitochondrial Genetics Group, School of Biomedicine, Faculty of Health and Medical Sciences, The University of Adelaide, Adelaide Health and Medical Sciences Building, Adelaide, SA 5000, Australia.
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