Beck Julia, Urnovitz Howard B, Riggert Joachim, Clerici Mario, Schütz Ekkehard
Chronix Biomedical GmbH, Goettingen, Germany.
Clin Chem. 2009 Apr;55(4):730-8. doi: 10.1373/clinchem.2008.113597. Epub 2009 Jan 30.
Circulating nucleic acids (CNAs) have been shown to have diagnostic utility in human diseases. The aim of this study was to sequence and organize CNAs to document typical profiles of circulating DNA in apparently healthy individuals.
Serum DNA from 51 apparently healthy humans was extracted, amplified, sequenced via pyrosequencing (454 Life Sciences/Roche Diagnostics), and categorized by (a) origin (human vs xenogeneic), (b) functionality (repeats, genes, coding or noncoding), and (c) chromosomal localization. CNA results were compared with genomic DNA controls (n = 4) that were subjected to the identical procedure.
We obtained 4.5 x 10(5) sequences (7.5 x 10(7) nucleotides), of which 87% were attributable to known database sequences. Of these sequences, 97% were genomic, and 3% were xenogeneic. CNAs and genomic DNA did not differ with respect to sequences attributable to repeats, genes, RNA, and protein-coding DNA sequences. CNA tended to have a higher proportion of short interspersed nuclear element sequences (P = 0.1), of which Alu sequences were significant (P < 0.01). CNAs had a significantly lower proportion of L1 and L2 long interspersed nuclear element sequences (P < 0.01). In addition, hepatitis B virus (HBV) genotype F sequences were found in an individual accidentally evaluated as a healthy control.
Comparison of CNAs with genomic DNA suggests that nonspecific DNA release is not the sole origin for CNAs. The CNA profiling of healthy individuals we have described, together with the detailed biometric analysis, provides the basis for future studies of patients with specific diseases. Furthermore, the detection of previously unknown HBV infection suggests the capability of this method to uncover occult infections.
循环核酸（CNA）已被证明在人类疾病诊断中具有实用价值。本研究的目的是对CNA进行测序和整理，以记录健康个体中循环DNA的典型特征。
提取51名健康个体的血清DNA，进行扩增，通过焦磷酸测序（454生命科学公司/罗氏诊断公司）进行测序，并按以下方式分类：（a）来源（人类与异种），（b）功能（重复序列、基因、编码或非编码），以及（c）染色体定位。将CNA结果与经过相同程序的基因组DNA对照（n = 4）进行比较。
我们获得了4.5×10⁵个序列（7.5×10⁷个核苷酸），其中87%可归因于已知数据库序列。在这些序列中，97%是基因组序列，3%是异种序列。CNA和基因组DNA在重复序列、基因、RNA和蛋白质编码DNA序列方面没有差异。CNA中短散在核元件序列的比例往往较高（P = 0.1），其中Alu序列具有显著性（P < 0.01）。CNA中L1和L2长散在核元件序列的比例显著较低（P < 0.01）。此外，在一名被意外评估为健康对照的个体中发现了乙型肝炎病毒（HBV）F基因型序列。
CNA与基因组DNA的比较表明，非特异性DNA释放不是CNA的唯一来源。我们所描述的健康个体的CNA谱，连同详细的生物特征分析，为未来对特定疾病患者的研究提供了基础。此外，对先前未知的HBV感染的检测表明该方法有发现隐匿感染的能力。
