Rapid detection of specific messenger RNAs in thyroid carcinomas by reverse transcription-PCR with degenerate primers: specific expression of oncofetal fibronectin messenger RNA in papillary carcinoma.
Rapid detection of specific messenger RNAs in thyroid carcinomas by reverse transcription-PCR with degenerate primers: specific expression of oncofetal fibronectin messenger RNA in papillary carcinoma.
复制标题
使用简并引物通过逆转录 PCR 快速检测甲状腺癌中的特异性信使 RNA：乳头状癌中癌胎纤连蛋白信使 RNA 的特异性表达。
作者：
Toru Takano;F. Matsuzuka;H. Sumizaki;Kanji Kuma;Nobuyuki Amino
寻找在癌症组织中特异性表达的mRNA对于基因诊断和治疗非常重要。然而，在人类癌症中发现这样的mRNA通常是非常困难的，这是因为从组织中可获得的RNA的体积有限以及RNA分析的许多技术困难。为了解决这些问题，本研究采用两种简并引物，通过逆转录PCR比较了甲状腺癌组织与正常和良性组织的mRNA。扩增的cDNA经非变性丙烯酰胺凝胶电泳分离后，选择癌组织中扩增出的3条条带，PCR扩增，克隆到T载体中。其中一条带经测序分析确定为癌胚纤连蛋白。用北方杂交和反转录-聚合酶链反应检测癌胚纤维连接蛋白mRNA在良、恶性肿瘤组织中的表达，发现其表达仅在乳头状癌和间变性癌中增高。因此，本方法快速检测癌组织中的特异性mRNA，其中之一，癌胚纤连蛋白mRNA，是乳头状癌基因诊断的良好靶点。
The search for mRNAs that are specifically expressed in cancer tissues is important for gene diagnosis and therapy. However, finding such mRNAs in human cancers is usually very difficult, both because of the limited volume of RNA obtainable from the tissues and the many technical difficulties of RNA analysis. To address these problems, the present study compared mRNA from thyroid cancer tissues with those from normal and benign tissues by reverse transcription-PCR using two degenerate primers. Amplified cDNAs were separated by electrophoresis with nondenaturing acrylamide gel, then three bands that are increased in cancer tissues were selected, reamplified by PCR, and cloned into T-vector. One of the bands was determined by sequencing analysis to be oncofetal fibronectin. The expression of oncofetal fibronectin mRNA in benign and malignant tissues was examined by Northern blot and reverse transcription-PCR using specific primers of its cDNA sequence, and its increased expression was observed only in papillary and anaplastic carcinomas. Thus, the present method rapidly detected specific mRNAs in cancer tissues, and one of these, oncofetal fibronectin mRNA, is a good target for gene diagnosis of papillary carcinoma.
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