The rapid advances in molecular genetics and reproductive technology during the past decade have led to the production of a large number of artificially produced animal genotypes. The synergy of these two fields has been especially influential in the mouse, since it is now the primary animal model for many human diseases. It is important, therefore, to reliably preserve the large numbers of mouse genotypes for use by future investigators as safely, simply, and economically as possible. Traditionally, mouse genotypes are stored by maintaining active breeding colonies of mutant mice. However, the cost of live colony maintenance has become prohibitively expensive. Alternatively, the banking of spermatozoa would be an efficient and cost effective approach for the storage of transgenic and mutant stocks. Several methods are available today for cryopreservation and maintenance of mouse sperm, but these require sophisticated freezing equipment and methodologies. More recently, freeze-drying for storage at ambient temperatures have been developed to circumvent some of the limitations of cryogenic storage. However, freeze-drying also relies on sophisticated and cumbersome equipment. Thus, there is still a pressing need to develop stable repositories of mouse sperm stored at ambient temperatures with simplicity for seamless dissemination. Evaporative drying offers a simpler, faster, and inexpensive approach that can be mastered quickly and accomplished successfully with minimal training. Furthermore, cellular stability is achieved quickly at ambient temperature without prolonged exposure to deleterious conditions during drying. Organisms that naturally enter a dried state during dormancy do so by evaporative drying, which suggests evaporative drying may be innocuous to cells under proper conditions.
In Specific Aim 1, we will optimize both the physicochemical and biological conditions to achieve high recovery of sperm after desiccation and long-term storage at ambient temperature.
In Specific Aim 2, we will develop engineered trehalose-derivatives that freely permeates the plasma membrane of the sperm for efficient loading of the protectant at amounts necessary for stabilization.
In Specific Aim 3, we will analyze the viability, health, and genetic and phenotypic fidelity of embryos and live born offspring generated from evaporatively dried sperm stored at ambient temperatures.
The rapid advances in molecular genetics and reproductive technology during the past decade have led to the production of a large number of artificially produced mouse genotypes that need to be stored reliably for use by future investigators. Evaporative drying offers a simple, fast, and inexpensive approach that can be mastered quickly and accomplished successfully with minimal training.
|Weng, Lindong; Tessier, Shannon N; Swei, Anisa et al. (2017) Controlled ice nucleation using freeze-dried Pseudomonas syringae encapsulated in alginate beads. Cryobiology 75:1-6|
|Li, Ming-Wen; Glass, Olivia C; Zarrabi, Jasmin et al. (2016) Cryorecovery of Mouse Sperm by Different IVF Methods Using MBCD and GSH. J Fertili In Vitro 4:|
|Liu, Jie; Tanrikut, Cigdem; Wright, Diane L et al. (2016) Cryopreservation of human spermatozoa with minimal non-permeable cryoprotectant. Cryobiology 73:162-7|
|Weng, Lindong; Tessier, Shannon N; Smith, Kyle et al. (2016) Bacterial Ice Nucleation in Monodisperse D2O and H2O-in-Oil Emulsions. Langmuir 32:9229-36|
|Abazari, Alireza; Meimetis, Labros G; Budin, Ghyslain et al. (2015) Engineered Trehalose Permeable to Mammalian Cells. PLoS One 10:e0130323|
|Liu, Jie; Lee, Gloria Y; Lawitts, Joel A et al. (2014) Live pups from evaporatively dried mouse sperm stored at ambient temperature for up to 2 years. PLoS One 9:e99809|
|Abazari, Alireza; Chakraborty, Nilay; Hand, Steven et al. (2014) A Raman microspectroscopy study of water and trehalose in spin-dried cells. Biophys J 107:2253-62|
|Li, Ming-Wen; Baridon, Brian; Trainor, Amanda et al. (2012) Mutant mice derived by ICSI of evaporatively dried spermatozoa exhibit expected phenotype. Reproduction 143:449-53|