Genetic and genomic architecture in eight strains of the laboratory opossum Monodelphis domestica

Abstract

The gray short-tailed opossum (Monodelphis domestica) is an established laboratory-bred marsupial model for biomedical research. It is a critical species for comparative genomics research, providing the pivotal phylogenetic outgroup for studies of derived vs ancestral states of genomic/epigenomic characteristics for eutherian mammal lineages. To characterize the current genetic profile of this laboratory marsupial, we examined 79 individuals from eight established laboratory strains. Double digest restriction site-associated DNA sequencing and whole-genome resequencing experiments were performed to investigate the genetic architecture in these strains. A total of 66,640 high-quality single nucleotide polymorphisms (SNPs) were identified. We analyzed SNP density, average heterozygosity, nucleotide diversity, and population differentiation parameter Fst within and between the eight strains. Principal component and population structure analysis clearly resolve the strains at the level of their ancestral founder populations, and the genetic architecture of these strains correctly reflects their breeding history. We confirmed the successful establishment of the first inbred laboratory opossum strain LSD (inbreeding coefficient F > 0.99) and a nearly inbred strain FD2M1 (0.98 < F < 0.99), each derived from a different ancestral background. These strains are suitable for various experimental protocols requiring controlled genetic backgrounds and for intercrosses and backcrosses that can generate offspring with informative SNPs for studying a variety of genetic and epigenetic processes. Together with recent advances in reproductive manipulation and CRISPR/Cas9 techniques for Monodelphis domestica, the existence of distinctive inbred strains will enable genome editing on different genetic backgrounds, greatly expanding the utility of this marsupial model for biomedical research.

Monodelphis domestica Induced Pluripotent Stem Cells Reveal Metatherian Pluripotency Architecture

“…To develop monoiPSC lines, neonatal skin fibroblast (neoSF) cultures from 10-day-old female pups from two fully inbred strains, FD2M1 ( F = 0.997) and LSD1 ( F = 0.995) [ 20 ] were established. While in log growth phase, cells were nucleofected with each of the purified OriP/EBNA episomal plasmids, pCE-hOCT3/4, pCE-hUL, pCE-hSK, pCXLE-hMLN, pCE-mp53DD, and pCXB-EBNA1, encoding human reprogramming factors OCT4 , SOX2 , NANOG , KLF4 , LIN28 and cMYC [ 21 , 22 ].…”

“…Primary neoSF cultures from two fully inbred M. domestica strains, FD2M1 and LSD1 [ 20 ], were established using the methodology previously developed in our laboratory [ 40 , 41 ]. The established primary neoSF cultures were expanded for 3–4 passages in MFGM containing Iscove’s Modified Dulbecco’s Medium (IMDM), 16% fetal bovine serum (FBS), 0.5% insulin, transferrin, selenite supplement (ITS g supplement), 2 mM L-glutamine, 1X Antibiotic-Antimycotic solution (all from Gibco, ThermoFisher Scientific, Waltham, MA, USA) and 2 mM CaCl 2 (Millipore Sigma, St. Louis, MO, USA).…”

Blood transcriptome responses to PFOA and GenX treatment in the marsupial biomedical model Monodelphis domestica

“…The gray short-tailed opossum, Monodelphis domestica , is the best marsupial model for biomedical research ( Fadem et al, 1982 ). This animal has been studied in a research laboratory environment for more than 40 years, has a well-assembled and well-annotated reference genome ( Mikkelsen et al, 2007 ), and includes two validated inbred strains that enable control of its genetic background ( Xiong et al, 2022 ). Marsupials (metatherian mammals) diverged from placentals (eutherian mammals) approximately 173–190 million years ago ( Woodburne et al, 2003 ; van Rheede et al, 2006 ; Luo et al, 2011 ).…”