Guo C, Van Damme B, Van Damme-Lombaerts R, Van den Berghe H, Cassiman J J, Marynen P
Center for Human Genetics, University of Leuven, Belgium.
Kidney Int. 1993 Dec;44(6):1316-21. doi: 10.1038/ki.1993.384.
PCR conditions were optimized to amplify the COL4A5 cDNA from lymphoblasts and kidney tissue. Sequencing of the COL4A5 mRNA isolated from the kidney of an Alport syndrome patient revealed two differences with the published sequence. One divergence, the insertion of an 18 bp sequence between exon 11 and 10 of the COL4A5 mRNA added two Gly-X-Y triplets to the COL4A5 sequence and was subsequently found in the mRNA of four normal kidney mRNA samples. This sequence was absent in all white blood cell RNA samples sequenced by us, indicating tissue specific splicing with the presence of an additional exon in kidney COL4A5 mRNA. This finding of differential splicing of COL4A5 mRNA in kidney and white blood cells might affect the use of white blood cell mRNA for the analysis of Alport mutations. Second, a complex mutation was detected in the mRNA from the AS patient introducing a premature stop codon in the message, deleting part of the triple helical domain and the complete NC domain. The mother of the patient was shown to be heterozygous for this mutation.
优化聚合酶链反应（PCR）条件以从淋巴母细胞和肾组织中扩增Ⅳ型胶原α5链（COL4A5）互补DNA（cDNA）。对从一名奥尔波特综合征患者肾脏中分离出的COL4A5信使核糖核酸（mRNA）进行测序，发现其与已发表序列存在两处差异。其中一处差异是，在COL4A5 mRNA的第11外显子和第10外显子之间插入了一段18碱基对（bp）的序列，这使得COL4A5序列中增加了两个甘氨酸-X-酪氨酸（Gly-X-Y）三联体，随后在四个正常肾脏mRNA样本的mRNA中也发现了该序列。在我们测序的所有白细胞RNA样本中均未发现该序列，这表明存在组织特异性剪接，即肾脏COL4A5 mRNA中存在一个额外的外显子。在肾脏和白细胞中COL4A5 mRNA的差异剪接这一发现，可能会影响利用白细胞mRNA分析奥尔波特突变的应用。其次，在该奥尔波特综合征（AS）患者的mRNA中检测到一个复杂突变，该突变在信息中引入了一个提前终止密码子，删除了部分三螺旋结构域和整个非胶原结构域（NC结构域）。结果显示，该患者的母亲为该突变的杂合子。
