Sloter E D, Lowe X, Moore II D H, Nath J, Wyrobek A J
Biology and Biotechnology Research Program, Lawrence Livermore National Laboratory, Livermore, CA 94550, USA.
Am J Hum Genet. 2000 Oct;67(4):862-72. doi: 10.1086/303088. Epub 2000 Aug 28.
Transmitted de novo structural chromosomal abnormalities, the majority of which are paternally derived, can lead to abnormal reproductive outcomes as well as genetic diseases in offspring. We developed and validated a new multicolor FISH procedure (sperm ACM, which utilizes DNA probes specific for the alpha [1cen], classical, [1q12], and midi [1p36.3] satellites of chromosome 1) which utilizes DNA probes specific for three regions of chromosome 1 to detect human sperm that carry numerical abnormalities plus two categories of structural aberrations: (1) duplications and deletions of 1pter and 1cen, and (2) chromosomal breaks within the 1cen-1q12 region. In healthy men, the average frequencies of sperm with duplications and deletions were (a) 4.5 +/- 0.5 and 4.1 +/- 1.3 per 10(4) involving 1pter and (b) 0.9 +/- 0.4 and 0.8 +/- 0.3 per 10(4) involving 1cen, respectively. The frequency of sperm exhibiting breaks within the 1cen-1q12 region was 14.1 +/- 1.2 per 10(4). Structural aberrations accounted for 71% of the abnormalities detected by sperm ACM, which was significantly higher than numerical abnormalities (P=2x10-8). Our findings also suggest that, for healthy men, (a) sperm carrying postmeiotic chromosomal breaks appear to be more prevalent than those carrying products of premeiotic or meiotic breakage or rearrangements, (b) the high frequency of chromosome breaks measured after "fertilization" by the hamster-egg cytogenetic method already appear to be present and detectable within human sperm by FISH, and (c) there are nonrandom and donor-specific distributions of breakpoint locations within 1q12 in sperm. FISH facilitates the analysis of much larger numbers of sperm than was possible when the hamster-egg method was used. Therefore, FISH-based procedures for simultaneously detecting chromosomal breaks, rearrangements, and numerical abnormalities in sperm may have widespread applications in human genetics, genetic toxicology, and reproductive medicine.
新发结构性染色体异常大多源自父方,可导致异常生殖结局以及子代患遗传病。我们开发并验证了一种新的多色荧光原位杂交技术(精子ACM,该技术利用针对1号染色体的α[1cen]、经典型[1q12]和中型[1p36.3]卫星区域的DNA探针),该技术利用针对1号染色体三个区域的DNA探针来检测携带数目异常以及两类结构畸变的人类精子:(1)1pter和1cen的重复与缺失,以及(2)1cen - 1q12区域内的染色体断裂。在健康男性中,涉及1pter的重复和缺失精子的平均频率分别为(a)每10⁴个中有4.5±0.5个和4.1±1.3个,以及(b)涉及1cen的每10⁴个中有0.9±0.4个和0.8±0.3个。在1cen - 1q12区域内出现断裂的精子频率为每10⁴个中有14.1±1.2个。结构畸变占精子ACM检测到的异常的71%,显著高于数目异常(P = 2×10⁻⁸)。我们的研究结果还表明,对于健康男性,(a)携带减数分裂后染色体断裂的精子似乎比携带减数分裂前或减数分裂期断裂或重排产物的精子更普遍,(b)通过仓鼠卵细胞遗传学方法在“受精”后测得的高频率染色体断裂似乎在人类精子中已经存在且可通过荧光原位杂交检测到,以及(c)精子中1q12内的断点位置存在非随机且供体特异性分布。与使用仓鼠卵方法时相比,荧光原位杂交技术有助于分析数量更多的精子。因此,基于荧光原位杂交技术同时检测精子中染色体断裂、重排和数目异常的方法可能在人类遗传学、遗传毒理学和生殖医学中有广泛应用。