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Title: RECENT IMPROVEMENTS IN EFFICIENCY OF FLOW CYTOMETRIC SORTING OF X AND Y- CHROMOSOME BEARING SPERM OF DOMESTIC ANIMALS: A REVIEW

Author
item GURNSEY, M - AGRESEARCH, NEW ZEALAND
item Johnson, Lawrence

Submitted to: Proceedings New Zealand Society of Animal Production
Publication Type: Review Article
Publication Acceptance Date: 5/10/1998
Publication Date: N/A
Citation: N/A

Interpretive Summary: Sexing of livestock semen for use in various practical circumstances in various areas of the world is now a virtual reality. The utilization of sexed semen in various livestock operations using artificial insemination has been enhanced due to the availability of recent improvements in sorting X and Y chromosome bearing sperm. In the studies described in this paper high speed sorting was employed in contrast to previous work where we were able to use only standard speed sperm sorting. The advantage of high speed has been to increase the sorting speed by five to ten times and thus make it economically feasible to use sexed semen with deep uterine artificial insemination in cattle and for in vitro fertilization in swine. These results will be utilized by organizations to develop the utility of sexing semen in practical applications.

Technical Abstract: In mammals the only method known to reliably separate X and Y sperm for producing offspring of a specific sex is flow cytometric sorting. This method is based on the observation that X-chromosome bearing sperm of domestic animals contain 3.5-4.2% more DNA than Y-chromosome bearing sperm. Relative DNA content is determined by quantitative staining with Hoechst 33342 and DNA content measured using a modified cell sorter. The major constraint to widespread application of this technology has been the slow sorting rate. Using modified standard speed cell sorters, sample flow rates average 2,000 sperm/sec with 25-35% orientation. Recently, a high speed sorter (MoFlo) modified for sperm sorting has been used and increases sorting rate by approximately 5-fold. Further, an orienting nozzle has been developed which increases the percentage of sperm that are orientated thus increasing the sort rate by a further 2 to 3-fold. Combined, these improvements have increased the production of sorted sperm from approximately 0.3 x 1,000,000/h with conventional sorters to at least 4 x 1,000,000/h with the high speed sorter. Preliminary testing suggests that purity of sort and sperm viability are not compromised by increased sorting rates. Pregnancies have been established in pigs and cows with sorted sperm from a modified MoFlo fitted with a novel nozzle. The improvements outlined here greatly enhance the Beltsville Sperm Sexing Technology and make it realistic to consider trials using sex sorted sperm for conventional AI as well as deep uterine AI in cattle.