title: Successful in vitro fertilization and generation of transgenics in Black and Tan Brachyury (BTBR) mice.
abstract: The Black and Tan Brachyury (BTBR) mouse strain is a valuable model for the study of long-term complications from obesity-induced type 2 diabetes mellitus and autism spectrum disorder. Due to technical difficulties with assisted reproduction, genetically modified animals on this background have previously been generated through extensive backcrossing, which is expensive and time-consuming. We successfully generated two separate transgenic mouse lines after direct zygote microinjection into this background strain. Additionally, we developed in vitro fertilization (IVF) methods for the BTBR mouse. We found low rates of fertilization and implantation in this strain, and identified the BTBR oocyte as the primary culprit of low success with BTBR IVF. We achieved an increase in live born pups from 5.9 to 35.6 % with IVF in the BTBR strain by use of BTBR females at a younger age (18-25 days), collection of oocytes 15-17 h after superovulation, and the use of supplemented fertilization media. This method eliminates the need for time consuming assisted embryo manipulations that are otherwise required for success with BTBR oocytes. This advancement provides an exciting opportunity to directly generate BTBR transgenics and gene-edited mice using both traditional and emerging genomic editing techniques, such as CRISPR/Cas9. These methods also allow effective colony preservation and rederivation with these strains. To our knowledge, this is the first report describing embryo manipulations in BTBR mice.
authors: Division of Endocrinology, Diabetes and Metabolism, Department of Medicine, University of Wisconsin-Madison, 4147 MFCB, 1685 Highland Ave, Madison, WI, 53705, USA.
authors: Mieke
authors: M
authors: Baan
authors: University of Wisconsin-Madison Biotechnology Center, 425 Henry Mall, Madison, WI, 53706, USA.
authors: Kathleen J
authors: KJ
authors: Krentz
authors: Division of Endocrinology, Diabetes and Metabolism, Department of Medicine, University of Wisconsin-Madison, 4147 MFCB, 1685 Highland Ave, Madison, WI, 53705, USA.
authors: Danielle A
authors: DA
authors: Fontaine
authors: Division of Endocrinology, Diabetes and Metabolism, Department of Medicine, University of Wisconsin-Madison, 4147 MFCB, 1685 Highland Ave, Madison, WI, 53705, USA. dbd@medicine.wisc.edu.|William S. Middleton Memorial Veterans Hospital, 2500 Overlook Terrace, Madison, WI, 53705, USA. dbd@medicine.wisc.edu.
authors: Dawn Belt
authors: DB
authors: Davis
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified / Q000235:genetics; D000067877:Autism Spectrum Disorder / Q000150:complications / Q000235:genetics / Q000473:pathology; D064112:Clustered Regularly Interspaced Short Palindromic Repeats / Q000235:genetics; D003924:Diabetes Mellitus, Type 2 / Q000150:complications / Q000235:genetics* / Q000473:pathology; D004195:Disease Models, Animal; D005307:Fertilization in Vitro / Q000379:methods*; D005326:Fetal Proteins / Q000235:genetics*; D006801:Humans; D051379:Mice; D009765:Obesity / Q000150:complications / Q000235:genetics* / Q000473:pathology; D009865:Oocytes / Q000254:growth & development; D020825:T-Box Domain Proteins / Q000235:genetics*; D000098908:Brachyury Protein
chemical_list: D005326:Fetal Proteins; D020825:T-Box Domain Proteins; D000098908:Brachyury Protein
keywords: Autism; BTBR; Diabetes mellitus; Embryo transfer; In vitro fertilization; Transgenic mouse model