遗传 ›› 2008, Vol. 30 ›› Issue (8): 983-990.doi: 10.3724/SP.J.1005.2008.00983

• 研究报告 • 上一篇    下一篇

引物原位标记技术快速检测人类染色体非整倍性

朱一剑; 刘涤石; 丁显平   

  1. 四川大学生命科学学院遗传医学研究所, 生物资源与生态环境教育部重点实验室, 成都 610064
  • 收稿日期:2007-12-16 修回日期:2008-03-12 出版日期:2008-08-10 发布日期:2008-08-10
  • 通讯作者: 丁显平

Detecting human chromosome anomalies with primed in situ label-ing (PRINS)

ZHU Yi-Jian; LIU Di-Shi; DING Xian-Ping   

  1. Key Laboratory of Bio-resources and Eco-environment of Ministry of Education, Institute of Medical Genetics, Sichuan University, Chengdu 610064, China
  • Received:2007-12-16 Revised:2008-03-12 Online:2008-08-10 Published:2008-08-10
  • Contact: DING Xian-Ping

摘要: 染色体数目异常是人类染色体疾病的重要类型, 经常导致胚胎丢失、胎儿流产、婴儿死亡、先天畸形和神经发育异常等出生缺陷。文章应用引物原位标记(Primed in situ labeling, PRINS)技术快速检测人类染色体非整倍性, 率先采用更新的非ddNTP阻断的多色PRINS技术, 对人类外周血淋巴细胞和精子等多种样本进行标记; 然后对不同靶标序列的标记效率及不同荧光色素的发光特点通过实验进行评估, 获得关于PRINS技术的多项反应原理参数, 并筛选标记顺序以获得均一稳定的标记效果, 最后进行临床FISH探针与PRINS的标记比较实验。通过实验比较PRINS技术与传统FISH技术之间的标记特点与差别, 评估PRINS的实际应用效果。在2.5 h内标记了同一精子核内的多条染色体, 单色以上标记达到99%。同时在人类外周血淋巴细胞中也得到较好的标记效果。与FISH技术相比, PRINS的这些优点使得它成为诊断染色体非整倍性变异的首选技术。

关键词: 非整倍性, 人类染色体, 引物原位标记

Abstract: Numerical chromosome anomaly was one of the most important kinds of human chromosme diseases by inducing pregnancy loss, miscarriage, infant death, congenital malformations and nerve damage. The present study was to establish a rapid, reliable and reasonable multicolor primed in situ labeling (PRINS) protocol for diagnosing numerical anomaly in human chromosome. First, nuclei of cultured lymphocytes and sperms were labeled with the method of PRINS, and then nuclei of cultured lymphocytes, sperms and other specimen were labeled with the method of updated non-ddNTP-blocking multicolor PRINS technique. The labeling effect of different target sequences and the feature of different fluorochromes were evaluated by experiment. Meanwhile, several parameters of PRINS were optimized to obtain more homogeneous and stable labeling effect. At last, the applicative value of PRINS was evaluated by comparing the clinical effect and labeling characteristics between FISH probe and PRINS. In the present study, several chromosomes were simultaneously marked successfully in the same sperm nucleus within 2.5 hours. And the frequency of one-color-labeling reached 99%. The many advantages, compared with FISH, make PRINS become the first choice in diagnosing diseases related to numerical anomaly in human chromosome.