Rath D, Long C R, Dobrinsky J R, Welch G R, Schreier L L, Johnson L A
Institute of Animal Science and Animal Behaviour, Mariensee, Neustadt, Germany.
J Anim Sci. 1999 Dec;77(12):3346-52. doi: 10.2527/1999.77123346x.
The objectives for the present experiments were to apply sperm sexing technology to an in vitro production system with porcine oocytes obtained from slaughterhouse material. On six experimental days, ovaries were obtained from an abattoir, and cumulus-oocyte-complexes were matured in vitro. Semen was collected from mature boars of proven fertility and was sorted for X-chromosome-bearing sperm, using the Beltsville Sperm Sexing Technology incorporating the use of high-speed sorting. A total of 5,378 oocytes were submitted for in vitro fertilization (IVF). Of these, 559 ova were stained for cytogenetic analysis 18 h after IVF. From the remaining 4,819 ova, 1,595 cleaved, and 1,300 of the cleaved embryos were transferred into 26 synchronized recipients (5 control gilts for unsorted sperm, 21 gilts for X-sorted sperm). In a test of two fertilization media (FERT-A vs FERT-B) higher cleavage rates (P<.05) were obtained when FERT-B was used as a fertilization medium for unsorted (43.4+/-5.1%) and sorted sperm (43.1+/-1.1%;), whereas in FERT-A unsorted sperm gave a cleavage rate of 17.9+/-4.4% and sorted sperm gave 30.4+/-1.4%. Additionally, cleavage rates were higher (P<.05) after fertilization with sorted sperm vs unsorted sperm, independent of fertilization medium. Cytogenetic analysis of ova revealed that more oocytes with unsorted than with sorted sperm remained in Metaphase 2 arrest (P<.05). This was also independent of the fertilization medium. Monospermic fertilization rates were the same for IVF with unsorted or sorted sperm, independent of the fertilization system, except FERT-A with unsorted sperm (P<.05). Polyspermic fertilization rates were highest in FERT-B (37.6+/-6.6). A total of 57 pigs were born from nine litters. Six litters from sexed sperm (X-sorted) produced 33 females (97%) and one male. Three litters from control transfers produced 23 pigs, 11 of which were female (48%). The sex ratio of the offspring was predicted based on the sort reanalysis of the sorted sperm for DNA content.
本实验的目的是将精子性别分选技术应用于体外生产系统，该系统使用从屠宰场材料中获取的猪卵母细胞。在六个实验日，从屠宰场获取卵巢，卵丘-卵母细胞复合体在体外成熟。从经证实具有生育能力的成年公猪采集精液，并使用结合高速分选的贝尔茨维尔精子性别分选技术对携带X染色体的精子进行分选。总共5378个卵母细胞用于体外受精（IVF）。其中，559个卵子在IVF后18小时进行细胞遗传学分析染色。从其余4819个卵子中，1595个发生分裂，1300个分裂胚胎被移植到26个同期受体中（5头对照母猪接受未分选精子，21头母猪接受X分选精子）。在对两种受精培养基（FERT-A与FERT-B）的测试中，当FERT-B用作未分选精子（43.4±5.1%）和分选精子（43.1±1.1%） 的受精培养基时，获得了更高的分裂率（P<0.05）；而在FERT-A中，未分选精子的分裂率为17.9±4.4%，分选精子的分裂率为30.4±1.4%。此外，无论受精培养基如何，分选精子受精后的分裂率均高于未分选精子（P<0.05）。卵子的细胞遗传学分析显示，未分选精子受精的卵子比分选精子受精的卵子更多地停留在中期2（P<0.05）。这也与受精培养基无关。单精子受精率在未分选或分选精子的IVF中相同，与受精系统无关，但FERT-A与未分选精子的情况除外（P<0.05）。多精子受精率在FERT-B中最高（37.6±6.6）。总共9窝产下57头仔猪。来自分选精子（X分选）的6窝产下33头雌性仔猪（97%）和1头雄性仔猪。来自对照移植的3窝产下23头仔猪，其中11头为雌性（48%）。根据对分选精子DNA含量的重新分析预测后代的性别比例。
