Johnson L A
Germplasm and Gamete Physiology Laboratory, Agricultural Research Service U. S. Department of Agriculture, Beltsville, Maryland, USA.
Dtsch Tierarztl Wochenschr. 1996 Aug-Sep;103(8-9):288-91.
Recent advances in the separation of X and Y chromosome bearing spermatozoa have led to the availability of a method (Beltsville Sperm Sexing Technology) to preselect the sex in several mammals. Progeny using this procedure have been produced in cattle, sheep, swine and laboratory animals. Mammalian sperm are inherently different in that the X sperm carries from 2.8 to 7.5% more DNA than the Y sperm. Individual sperm DNA can be determined and used as the differentiating characteristic with flow cytometry and cell sorting instrumentation especially modified to measure small amounts of DNA in sperm. The process utilizes the fluorochrome Hoechst 33342 to bind to the DNA. The relative DNA is measured by passing the living sperm through a laser beam and collecting the light energy from the individual sperm. Data is acquired and used to select the particular sperm for deflection into collection tubes. The proportions of sorted X and Y sperm in each tube can be validated by reanalyzing an aliquot for DNA content. This value is then used to predict the outcome of fertilization and subsequent gestation. The sorted sperm are used to inseminate eggs via in vitro fertilization (IVF) or by surgical insemination into the oviduct or the uterus of appropriate females. Sperm are sorted at the rate of 0.5 million per hour for most species with the expectation of 90% or greater of one sex or the other being born. Progeny in cattle using IVF have been produced at greater than 90% accuracy. Rabbits have produced greater than 90% females using this process. Progeny produced from pigs average 85% for one sex or the other. All progeny produced (N = or > 300) have exhibited completely normal morphological appearance and normal reproductive function. Because of the inability to obtain large numbers of sorted sperm in a short amount of time, the technologies use for regular artificial insemination would not be practical in most domestic species. This sexing technology however is very applicable where IVF, intrauterine or intratubal insemination are convenient means for producing offspring. In addition, the recent advent of ultrasound guided insemination in cattle may provide and opportunity to use this technology for much lower numbers of sperm per insemination than previously thought possible. Using less than 2 x 10 sorted X or Y sperm would move the technology one step closer to practicality.
在携带X和Y染色体精子的分离方面取得的最新进展，已催生出一种在多种哺乳动物中进行性别预选的方法（贝尔茨维尔精子性别分选技术）。利用这一程序已培育出了牛、羊、猪和实验动物的后代。哺乳动物的精子本质上存在差异，即X精子携带的DNA比Y精子多2.8%至7.5%。可以测定单个精子的DNA，并将其用作区分特征，借助特别改装用于测量精子中少量DNA的流式细胞仪和细胞分选仪器来实现。该过程利用荧光染料Hoechst 33342与DNA结合。通过让活精子穿过激光束并收集单个精子的光能来测量相对DNA含量。获取数据并用于选择特定的精子，使其偏转到收集管中。每个管中分选的X和Y精子的比例可通过重新分析一份DNA含量的等分试样来验证。然后使用该值来预测受精和随后妊娠的结果。分选后的精子通过体外受精（IVF）或通过手术授精到合适雌性动物的输卵管或子宫中来使卵子受精。对于大多数物种而言，精子的分选速度为每小时50万个，预期出生的后代中某一性别的比例达到90%或更高。使用IVF技术培育的牛后代的准确率超过90%。使用这一过程培育出的兔子中，雌性比例超过90%。猪的后代中某一性别的平均比例为85%。所有培育出的后代（N = 或> 300）均表现出完全正常的形态外观和正常的生殖功能。由于无法在短时间内获得大量分选后的精子，常规人工授精所使用的技术在大多数家养物种中并不实用。然而，这种性别分选技术在IVF、子宫内或输卵管内授精是方便的后代生产方式的情况下非常适用。此外，牛中最近出现的超声引导授精可能提供一个机会，使得每次授精使用的精子数量比以前认为可能的数量少得多时也能应用该技术。使用少于2×10个分选后的X或Y精子将使该技术向实用性迈进一大步。
