目的·采用全外显子测序技术筛查完全性肺静脉异位引流（total anomalous pulmonary venous connection，TAPVC）的可能致病基因，并对其功能进行验证。方法·选择2014—2019年在上海交通大学医学院附属新华医院及上海儿童医学中心确诊的100例TAPVC患儿（患儿组）以及120例健康儿童（对照组）为研究对象。收集2组儿童的血液样本，抽提全血基因组DNA并行外显子测序，以筛查TAPVC的潜在致病基因。通过Mutation Taster、SIFT、PolyPhen-2网站筛选致病基因的有害突变位点，并行Sanger测序。构建PDX1野生型（野生组）及突变型（突变组）质粒，转染入HUVEC细胞后，分别采用实时荧光定量PCR（quantitative real-time PCR，qPCR）和蛋白质印迹法检测突变对PDX1的mRNA和蛋白水平的影响。采用STRING数据库进行蛋白与蛋白之间的相互作用分析，并采用qPCR研究PDX1调节的下游基因的表达。结果·在TAPVC患儿中发现致病基因为PDX1，Sanger测序显示该基因存在2个新发突变，即c.C237A （P33T）和c.C237G（P33A）。与野生组相比，2个突变组（CA组、CG组）的PDX1 mRNA水平没有显著变化，但蛋白相对表达量有明显增加，即分别是野生组的2.9倍和3.4倍（P=0.000，P=0.001）。蛋白质相互作用分析的结果显示，PDX1与SOX17相关联；且qPCR结果显示，PDX1过表达可下调HUVEC细胞中SOX17的表达。结论·PDX1的2个新发错义突变可影响其转录后翻译，且PDX1可能是通过调控SOX17来参与TAPVC的发生与发展。

Objective ·To explore the possible pathogenic gene of total anomalous pulmonary venous connection (TAPVC) by whole exon sequencing and verify its function. Methods ·One hundred TAPVC children (case group) and one hundred and twenty healthy children (control group) in Xinhua Hospital and Shanghai Children's Medical Center, Shanghai Jiao Tong University School of Medicine from 2014 to 2019 were included. The blood samples from the two groups of children were collected, and whole blood genomic DNA was extracted for exon sequencing to screen out the potential pathogenic genes of TAPVC. Harmful mutation sites of pathogenic genes were screened through Mutation Taster, SIFT and PolyPhen-2 websites, and then conducted by Sanger sequencing. The wild-type (wild-type group) and mutant (mutant group) plasmids of PDX1 were transfected into HUVEC cells. Quantitative real-time PCR (qPCR) and Western blotting were used to detect the effects of mutations on mRNA and protein levels of PDX1, respectively. The STRING database was used to analyze the interaction between proteins, and qPCR was used to determine the expressions of downstream genes regulated by PDX1. Results ·Pathogenic PDX1 was found in TAPVC children, and Sanger sequencing revealed two novel variants in the gene: c.C237A (P33T) and c. C237G (P33A). Compared with the wild-type group, there was no significant difference in PDX1 mRNA levels in the two mutant groups, but there was a significant increase in relative protein expression of the CA group and CG group, which was 2.9 and 3.4 times higher than the wild-type group, respectively (P=0.000, P=0.001). Protein interaction analysis demonstrated that PDX1 was associated with SOX17. qPCR results showed that overexpression of PDX1 could downregulate the expression of SOX17 in HUVEC. Conclusion ·The two novel PDX1 missense mutations can affect the process of PDX1 post-transcriptional translation, indicating that PDX1 may participate in the occurrence and development of TAPVC by regulating SOX17.