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Transgenic Research

, Volume 22, Issue 4, pp 873–875 | Cite as

Improvement of pronuclear injection-based targeted transgenesis (PITT) by iCre mRNA-mediated site-specific recombination

  • Masato OhtsukaEmail author
  • Hiromi Miura
  • Hideki Hayashi
  • Hirofumi Nakaoka
  • Minoru Kimura
  • Masahiro Sato
  • Channabasavaiah B. Gurumurthy
  • Hidetoshi Inoko
Technical Update

Keywords

Generation Vector Transgenic Rate Developmental Arrest Supplementary Movie Donor Vector 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Acknowledgments

We are grateful to K. Yamagata for pcDNA3.1EGFP-poly(A83) (Yamagata et al. 2005), and I. M. Verma for iCre plasmid (Addgene plasmid 12336). We also thank S. Ogiwara and T. Sato for technical assistance. We are grateful to Manju George for critical reading of the manuscript and for English corrections. This work is supported by the Grant-in-Aid for Young Scientists (B) (20700368) from the Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan, by Takeda Science Foundation, by the A-STEP (Adaptable and Seamless Technology Transfer Program through Target-driven R&D) from Japan Science and Technology Agency (JST), and by 2011–2012 Tokai University School of Medicine, Project Research to M.O.

Supplementary material

11248_2013_9703_MOESM1_ESM.docx (55 kb)
Supplementary material 1 (DOCX 54 kb)

This movie file shows injections of donor vector DNA/iCre mRNA mixture into the zygotes according to the Method 1 (see “Supplementary materials and methods”). These injections are demonstration using BDF2 zygotes (MPG 4624 kb)

This movie file shows injections of donor vector DNA/iCre mRNA mixture into the zygotes according to the Method 2 (see “Supplementary materials and methods”). These injections are demonstration using BDF2 zygotes (MPG 3048 kb)

References

  1. Fielder TJ, Montoliu L (2011) Transgenic production benchmarks. In: Advanced protocols for animal transgenesis : an ISTT manual. Springer Protocols Handbooks. Springer, pp 81–97. doi: 10.1007/978-3-642-20792-1_5
  2. Ohtsuka M, Ogiwara S, Miura H, Mizutani A, Warita T, Sato M, Imai K, Hozumi K, Sato T, Tanaka M, Kimura M, Inoko H (2010) Pronuclear injection-based mouse targeted transgenesis for reproducible and highly efficient transgene expression. Nucleic Acids Res 38:e198PubMedCrossRefGoogle Scholar
  3. Ohtsuka M, Miura H, Nakaoka H, Kimura M, Sato M, Inoko H (2012a) Targeted transgenesis through pronuclear injection of improved vectors into in vitro fertilized eggs. Transgenic Res 21:225–226PubMedCrossRefGoogle Scholar
  4. Ohtsuka M, Miura H, Sato M, Kimura M, Inoko H, Gurumurthy CB (2012b) PITT: pronuclear injection-based targeted transgenesis, a reliable transgene expression method in mice. Exp Anim 61:489–502PubMedCrossRefGoogle Scholar
  5. Shimshek DR, Kim J, Hubner MR, Spergel DJ, Buchholz F, Casanova E, Stewart AF, Seeburg PH, Sprengel R (2002) Codon-improved Cre recombinase (iCre) expression in the mouse. Genesis 32:19–26PubMedCrossRefGoogle Scholar
  6. Yamagata K, Yamazaki T, Yamashita M, Hara Y, Ogonuki N, Ogura A (2005) Noninvasive visualization of molecular events in the mammalian zygote. Genesis 43:71–79Google Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2013

Authors and Affiliations

  • Masato Ohtsuka
    • 1
    Email author
  • Hiromi Miura
    • 1
  • Hideki Hayashi
    • 5
  • Hirofumi Nakaoka
    • 2
  • Minoru Kimura
    • 1
  • Masahiro Sato
    • 3
  • Channabasavaiah B. Gurumurthy
    • 4
  • Hidetoshi Inoko
    • 1
  1. 1.Division of Basic Medical Science and Molecular Medicine, Department of Molecular Life ScienceTokai University School of MedicineIseharaJapan
  2. 2.Division of Human Genetics, Department of Integrated GeneticsNational Institute of GeneticsMishimaJapan
  3. 3.Section of Gene Expression Regulation, Frontier Science Research CenterKagoshima UniversityKagoshimaJapan
  4. 4.Department of Genetics, Cell Biology and AnatomyUniversity of Nebraska Medical CenterOmahaUSA
  5. 5.Education and Research Support CenterTokai UniversityIseharaJapan

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