Engineering Research Centre of Molecular Diagnostics, Ministry of Education, Department of Biomedical Sciences and the Key Laboratory of the Ministry of Education for Cell Biology and Tumor Cell Engineering, School of Life Sciences, Xiamen University, Xiamen, China.
Clin Chem. 2010 Sep;56(9):1451-9. doi: 10.1373/clinchem.2010.146472. Epub 2010 Jul 9.
Trisomies 13, 18, and 21 account for the majority of chromosomal aneuploidies detected in prenatal diagnosis. Diagnosis of these trisomies relies mainly on karyotype analysis. Several molecular methods have been developed for trisomy detection, but performance or throughput limitations of these methods currently constrain their use in routine testing.
We developed multiplex ligation-dependent probe amplification-based real-time PCR (MLPA/rtPCR) to simultaneously detect these 3 trisomy conditions with a single reaction. We applied the method to DNA isolated from 144 blinded clinical samples that included 32 cases of trisomy 21, 11 cases of trisomy 18, 1 case of trisomy 13, and 100 unaffected control samples; results were compared with karyotype analysis.
As judged by the results of the karyotype analysis, MLPA/rtPCR correctly detected all 44 cases of trisomy in the analysis of the blinded clinical samples. The method was able to detect a change in chromosome dosage as low as 1.2-fold.
This novel PCR-based technology simultaneously identified 3 types of trisomy in a single reaction and accurately detected trisomy with mosaicism, while reducing assay times and costs compared with conventional methods. The MLPA/rtPCR approach may have applicability in noninvasive prenatal diagnosis with maternal blood samples.
在产前诊断中，三体 13、18 和 21 占染色体非整倍体的大多数。这些三体的诊断主要依赖于核型分析。已经开发了几种用于三体检测的分子方法，但这些方法的性能或通量限制目前限制了它们在常规检测中的应用。
我们开发了基于多重连接依赖性探针扩增的实时 PCR（MLPA/rtPCR），可通过单次反应同时检测这 3 种三体情况。我们将该方法应用于从 144 个盲法临床样本中提取的 DNA，这些样本包括 32 例 21 三体、11 例 18 三体、1 例 13 三体和 100 例正常对照样本；结果与核型分析进行了比较。
根据核型分析结果判断，MLPA/rtPCR 在对盲法临床样本的分析中正确检测到了 44 例三体。该方法能够检测到染色体剂量变化低至 1.2 倍。
这种新的基于 PCR 的技术可在单次反应中同时鉴定 3 种三体，并准确检测嵌合体三体，同时与传统方法相比，缩短了检测时间和成本。MLPA/rtPCR 方法可能适用于非侵入性产前诊断中的母体外周血样本。
