Non-invasive ovulation tracking enables genetic engineering in wild rodents

Joanna Buchthal; Emma J Chory; Zachary Hill; Christy Dennison; Boqiang Tu; Rick P Wierenga; Çağrı Çevrim; Stefan Golas; Gabriel Meier; Tammy C T Lan; Hattie Chung; Magalie Boucher; Sam R Telford; Kara L McKinley; Styliani Markoulaki; Rudolf Jaenisch; Kevin M Esvelt

doi:10.1016/j.crmeth.2026.101311

Non-invasive ovulation tracking enables genetic engineering in wild rodents

Cell Rep Methods. 2026 Feb 23;6(2):101311. doi: 10.1016/j.crmeth.2026.101311. Epub 2026 Feb 17.

Authors

Joanna Buchthal¹, Emma J Chory², Zachary Hill³, Christy Dennison³, Boqiang Tu⁴, Rick P Wierenga³, Çağrı Çevrim⁵, Stefan Golas³, Gabriel Meier⁶, Tammy C T Lan⁷, Hattie Chung⁸, Magalie Boucher⁹, Sam R Telford¹⁰, Kara L McKinley¹¹, Styliani Markoulaki¹², Rudolf Jaenisch¹³, Kevin M Esvelt¹⁴

Affiliations

¹ Media Laboratory, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Mice Against Ticks, Lincoln, MA 01773, USA. Electronic address: buchthal@mit.edu.
² Media Laboratory, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Biomedical Engineering, Duke University, Durham, NC 27705, USA.
³ Media Laboratory, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
⁴ Media Laboratory, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
⁵ Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02138, USA.
⁶ Whitehead Institute for Biomedical Research, Cambridge, MA 02142, USA; RNA Biology and Molecular Physiology, Faculty for Biology, Bielefeld University, Bielefeld, Germany.
⁷ Broad Institute of Harvard and MIT, Cambridge, MA 02142, USA; Department of Molecular and Cellular Biology, Harvard University, Boston, MA, USA.
⁸ Yale University School of Medicine, New Haven, CT 06520, USA.
⁹ Division of Comparative Medicine, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
¹⁰ Department of Infectious Disease and Global Health, Tufts University, Grafton, MA 01536, USA.
¹¹ Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02138, USA; Howard Hughes Medical Institute, Cambridge, MA 02138, USA.
¹² Whitehead Institute for Biomedical Research, Cambridge, MA 02142, USA.
¹³ Whitehead Institute for Biomedical Research, Cambridge, MA 02142, USA; Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
¹⁴ Media Laboratory, Massachusetts Institute of Technology, Cambridge, MA 02139, USA. Electronic address: esvelt@mit.edu.

Abstract

Many non-model rodent species are inaccessible to genetic engineering due to our limited understanding of their reproductive biology. Here, we present a low-cost, camera-based estrous-tracking technology that enables transgenesis in the white-footed mouse Peromyscus leucopus, a key reservoir for Lyme disease. We demonstrate the efficient generation of pregnant and pseudopregnant mice via timed ovulation, provide protocols for embryo generation, cultivation, microinjection, and transplantation as well as an accurate developmental timeline, and report the first engineered Peromyscus. The same technology successfully tracked conserved estrous-linked cycling behavior in other rodents, including hamsters. Finally, estrous tracking differentiated reproductively healthy, geriatric female Peromyscus from those with declining fertility based solely on their activity, providing a non-invasive method for studying reproductive senescence. Collectively, these tools represent a critical resource for engineering non-model rodents, advance the long-lived Peromyscus as a model organism, and will prove essential to heritably immunizing wild rodent populations against Lyme disease.

Keywords: CP: developmental biology; CP: genetics; CRISPR-Cas9 transgenesis; Lyme disease reservoir; Peromyscus leucopus; automated activity monitoring; estrous cycle tracking; machine learning in cytology; non-invasive phenotyping; non-model rodents; reproductive senescence; white-footed mouse.

MeSH terms

Animals
Animals, Wild
Female
Genetic Engineering* / methods
Male
Mice
Ovulation* / genetics
Ovulation* / physiology
Peromyscus* / genetics
Peromyscus* / physiology
Pregnancy