Optimized toolkit for the manipulation of immortalized axolotl fibroblasts
The axolotl salamander model has broad utility for regeneration studies, but this model is limited by a lack of efficient cell-culture-based tools. The Axolotl Limb-1 (AL-1) fibroblast line, the only available immortalized axolotl cell line, was first published over 20 years ago, but many established molecular biology techniques, such as lipofectamine transfection, CRISPR-Cas9 mutagenesis, and antibiotic selection, work poorly or remain untested in AL-1 cells. Innovating technologies to manipulate AL-1 cells in culture and study their behavior following transplantation into the axolotl will complement in-vivo studies, decrease the number of animals used, and enable the faster, more streamlined investigation of regenerative biology questions. Here, we establish transfection, mutagenesis, antibiotic selection, and in-vivo transplantation techniques in axolotl AL-1 cells. These techniques will enable efficient culture with AL-1 cells and guide future tool development for the culture and manipulation of other salamander cell lines.
Authors
- Paul Gilbert
- Steven J. Blair (ORCID: https://orcid.org/0000-0002-4297-9363)
- Ryan T. Kim (ORCID: https://orcid.org/0009-0007-3920-4982)
- Konstantinos Sousounis (ORCID: https://orcid.org/0000-0002-5577-6207)
- S. Loeffler Jay
- Sebastian Böhm (ORCID: https://orcid.org/0009-0006-4599-6309)
- Elly M. Tanaka (ORCID: https://orcid.org/0000-0003-4240-2158)
- Nathan R. Souchet (ORCID: https://orcid.org/0000-0002-1255-4268)
- Jihee Han (ORCID: https://orcid.org/0000-0002-4220-5050)
- Sangwon Min (ORCID: https://orcid.org/0000-0001-9744-5847)
- Jessica L. Whited (ORCID: https://orcid.org/0000-0002-3709-6515)
- Ji‐Feng Fei (ORCID: https://orcid.org/0000-0003-2426-9402)
- Duygu Payzin‐Dogru (ORCID: https://orcid.org/0000-0002-3717-0244)
- Benjamin Tajer (ORCID: https://orcid.org/0000-0001-7791-1249)
- Aaron M. Savage (ORCID: https://orcid.org/0000-0002-2636-6145)
- Stéphane Roy (ORCID: https://orcid.org/0000-0002-4504-0968)
- M. Kidd (ORCID: https://orcid.org/0009-0005-6770-0866)
- Noah J. Lopez
- Anna G. Luong
- Hani D. Singer (ORCID: https://orcid.org/0009-0001-7722-2758)
- Eric Wynn
- Glory Kalu
- Tim Froitzheim
- J. Arly Nelson
- Noora Harake (ORCID: https://orcid.org/0009-0008-6166-1247)
- Alparslan Karabacak
- Katherine Courtemanche
- Antoine Decaux
Institutions
- Université Claude Bernard Lyon 1 (FR)
- École Normale Supérieure de Lyon (FR)
- Institute of Molecular Biotechnology (AT)
- Research Institute of Molecular Pathology (AT)
- Harvard University Press (US)
- Vienna Biocenter (AT)
- Guangdong Academy of Medical Sciences (CN)
- Guangdong Provincial People's Hospital (CN)
- Harvard Stem Cell Institute (US)
- Southern Medical University (CN)
- Université de Montréal (CA)
Publication Details
- Journal
- Methods
- Published
- 2025-04-05
- DOI
- https://doi.org/10.1016/j.ymeth.2025.03.019
- Primary Topic
- RNA Interference and Gene Delivery
- Type
- article
- Field-Weighted Citation Impact
- 0.00