Conserving, Distributing and Managing Genetically Modified Mouse Lines by Sperm Cryopreservation - Genetic Engineering: Recent Developments in Applications - page 63

oocytes during transit through the female reproductive tract. As reviewed by Vis-
conti et al[7], sperm fertilization ability is associated with plasma membrane re-
organization and increases in intracellular calcium levels and in Reactive Oxygen
Species[8] (ROS). Because cryopreservation modifies aspects of sperm function
associated with fertilization capacity[9], perhaps these processes can be modulated
to increase the fertility of cryopreserved mouse sperm. Thus, the objective of this
work was to develop economical processes to cryopreserve C57BL/6J sperm that
retain or enhance fertilizing capacity.
Table 1. Dependency of impaired fertility on the genetic background of cryopreserved mouse sperm.
Because variable cooling and warming rates have been observed with some
methods[10], our effort began by defining reproducible processes for cryopre-
serving and thawing mouse sperm. Our methods were then refined to enhance
the ability of cryopreserved C57BL/6J sperm to fertilize C57BL/6J oocytes. The
overall effectiveness of our approach as a tool for the routine management of GM
mouse lines, was demonstrated by cryopreserving and recovering 735 GM lines
maintained on twelve genetic backgrounds, including 527 GM lines maintained
on the C57BL/6J background.
results and discussion
Definition of Cooling and Warming Rates
More C57BL/6J sperm retained intact membranes with moderate cooling
(37±1°C/min) than with rapid cooling (94±2°C/min; Table 2). This is in accord
with previous work[10], [11]. Importantly, using the device and process reported