Lin Y C, Canatan H, Chang C J, Hu Y F, Chen R, Yu C Y, Brueggemeier R W, Somers W J
Department of Veterinary Physiology and Pharmacology, College of Veterinary Medicine, Ohio State University, Columbus 43210-1092.
J Med. 1994;25(1-2):41-64.
This study examined the expression of keratinocyte growth factor (KGF) gene in human and canine prostatic tissues. KGF transcript was detected in normal human prostatic tissues by reverse transcription and polymerase chain reactions (RT-PCR). PCR-generated human KGF complementary DNA (cDNA) clone was confirmed by restriction enzyme digestion analysis and partial DNA sequencing. Expression of KGF in archival canine benign prostatic hyperplastic tissues was also examined. In a pilot experiment, RNAs isolated from formalin-fixed (FF) and formalin-fixed and paraffin-embedded (FFPE) canine prostatic tissues were shown to be of sufficient quality to permit amplification of KGF mRNA by RT-PCR. The transcript of a housekeeping gene, glucose-6-phosphate dehydrogenase (G6PD), was detected by RT-PCR indicating the quality of RNAs to be more than adequate for RNA expression analysis. Later, total RNA from two archival canine FF prostate tissue types, benign prostatic hyperplasis and mild glandular hyperplasia, were used to amplify canine KGF transcripts. Southern hybridization analysis using rat and human KGF cDNAs as probes confirmed the fidelity of the amplified PCR product and it was indeed canine KGF.
本研究检测了人及犬前列腺组织中角质形成细胞生长因子（KGF）基因的表达。通过逆转录聚合酶链反应（RT-PCR）在正常人前列腺组织中检测到KGF转录本。通过限制性内切酶消化分析和部分DNA测序对PCR扩增得到的人KGF互补DNA（cDNA）克隆进行了确认。同时也检测了KGF在存档的犬良性前列腺增生组织中的表达。在一项预实验中，从福尔马林固定（FF）以及福尔马林固定石蜡包埋（FFPE）的犬前列腺组织中分离得到的RNA，经证明具有足够的质量，可用于通过RT-PCR扩增KGF mRNA。通过RT-PCR检测看家基因葡萄糖-6-磷酸脱氢酶（G6PD）的转录本，表明RNA的质量足以进行RNA表达分析。随后，使用来自两种存档的犬FF前列腺组织类型（良性前列腺增生和轻度腺性增生）的总RNA来扩增犬KGF转录本。以大鼠和人KGF cDNA作为探针进行的Southern杂交分析证实了扩增的PCR产物的准确性，其确实为犬KGF。
