Superovulation Using the Combined Administration of Inhibin Antiserum and Equine Chorionic Gonadotropin Increases the Number of Ovulated Oocytes in C57BL/6 Female Mice

Abstract

Superovulation is a reproductive technique generally used to produce genetically engineered mice. Superovulation in mice involves the administration of equine chorionic gonadotropin (eCG) to promote follicle growth and then that of human chorionic gonadotropin (hCG) to induce ovulation. Previously, some published studies reported that inhibin antiserum (IAS) increased the number of ovulated oocytes in ddY and wild-derived strains of mice. However, the effect of IAS on the C57BL/6 strain, which is the most widely used inbred strain for the production of genetically engineered mice, has not been investigated. In addition, the combined effect of IAS and eCG (IASe) on the number of ovulated oocytes in superovulation treatment has not been examined. In this study, we examined the effect of IAS and eCG on the number of ovulated oocytes in immature female mice of the C57BL/6 strain in superovulation treatment. Furthermore, we evaluated the quality of obtained oocytes produced by superovulation using IASe by in vitro fertilization (IVF) with sperm from C57BL/6 or genetically engineered mice. The developmental ability of fresh or cryopreserved embryos was examined by embryo transfer. The administration of IAS or eCG had a similar effect on the number of ovulated oocytes in C57BL/6 female mice. The number of ovulated oocytes increased to about 3-fold by the administration of IASe than by the administration of IAS or eCG alone. Oocytes derived from superovulation using IASe normally developed into 2-cell embryos by IVF using sperm from C57BL/6 mice. Fresh or cryopreserved 2-cell embryos produced by IVF between oocytes of C57BL/6 mice and sperm from genetically engineered mice normally developed into live pups following embryo transfer. In summary, a novel technique of superovulation using IASe is extremely useful for producing a great number of oocytes and offspring from genetically engineered mice.

Pharmacological and genetic reappraisals of protease and oxidative stress pathways in a mouse model of obstructive lung diseases

“…The mice were genotyped by manufacturer’s recommendation with the following primer sets; FW 5′-CTTCCAAGAGTTCAACTACCG-3′ and RV 5′-TCTACCAGCTCAGCCACAGTG-3′ (245 bp). In some experiments, reproductive techniques established by Center for Animal Resources and Development (CARD) were applied to efficiently produce an optimal number of mice with stably exhibiting pulmonary phenotypes61. The mice were housed in a vivarium in accordance with the guidelines of the animal facility center of Kumamoto University and were fed with normal chow ad libitum .…”

Culture time of vitrified/warmed zygotes before microinjection affects the production efficiency of CRISPR-Cas9-mediated knock-in mice

“…Recently, we have developed an ultra-superovulation method to collect more oocytes from female mice compared with conventional superovulation methods (Takeo and Nakagata, 2015), and we demonstrated that zygotes created via ultra-superovulation combined with IVF are also applicable for the production of various GM mice by direct microinjection of genome editing reagents (Nakagawa et al, 2016). …”

High-Speed Mouse Backcrossing Through the Female Germ Line

“…Co-injection of inhibin antiserum and PMS, also termed hyperovulation, is effective in several wild-derived mouse strains and has recently been shown to increase superovulation yields approximately 3-fold in C57BL/6J mice [27]. This method may therefore increase the feasibility of high-speed backcrossing onto a C57BL/6J background and potentially other strains, by providing a larger number of offspring to screen, at the cost of transiently increasing colony size and number of animals involved.…”