Gene Expression Studies in Formalin-Fixed Paraffin-Embedded Samples of Cutaneous Cancer: The Need for Reference Genes

被引：3
作者：
Garcia-Perez, Omar [1 ,2 ,3 ]
Melgar-Vilaplana, Leticia [4 ]
Cordoba-Lanus, Elizabeth [1 ,2 ,3 ]
Fernandez-de-Misa, Ricardo [1 ,2 ,5 ]
机构：
[1] Hosp Univ Nuestra Senora Candelaria, Res Unit, Ctra Gral Rosario 145, Santa Cruz de Tenerife 38010, Spain
[2] Univ Laguna, Calle Padre Herrera, San Cristobal de La Lagu 38200, Spain
[3] Inst Univ Enfermedades Trop Salud Publ Canarias I, San Cristobal de La Lagu 38296, Spain
[4] Hosp Univ Nuestra Senora Candelaria, Dept Pathol, Ctra Gral Rosario 145, Santa Cruz de Tenerife 38010, Spain
[5] Hosp Univ Nuestra Senora Candelaria, Dept Dermatol, Ctra Gral Rosario 145, Santa Cruz de Tenerife 38010, Spain
来源：
CURRENT ISSUES IN MOLECULAR BIOLOGY | 2021年 / 43卷 / 03期
关键词：
reference gene; malignant melanoma; cutaneous squamous cell carcinoma; qPCR; formalin-fixed paraffin-embedded; REAL-TIME PCR; HOUSEKEEPING GENES; VALIDATION; RNA; NORMALIZATION; BIOMARKER; SELECTION;
D O I：
10.3390/cimb43030151
中图分类号：
Q5 [生物化学]; Q7 [分子生物学];
学科分类号：
071010 ; 081704 ;
摘要：
Formalin-fixed paraffin-embedded (FFPE) tumour samples may provide crucial data regarding biomarkers for neoplasm progression. Analysis of gene expression is frequently used for this purpose. Therefore, mRNA expression needs to be normalized through comparison to reference genes. In this study, we establish which of the usually reported reference genes is the most reliable one in cutaneous malignant melanoma (MM) and cutaneous squamous cell carcinoma (CSCC). ACTB, TFRC, HPRT1 and TBP expression was quantified in 123 FFPE samples (74 MM and 49 CSCC biopsies) using qPCR. Expression stability was analysed by NormFinder and Bestkeeper softwares, and the direct comparison method between means and SD. The in-silico analysis with BestKeeper indicated that HPRT1 was more stable than ACTB and TFRC in MM (1.85 vs. 2.15) and CSCC tissues (2.09 vs. 2.33). The best option to NormFinder was ACTB gene (0.56) in MM and TFRC (0.26) in CSCC. The direct comparison method showed lower SD means of ACTB expression in MM (1.17) and TFRC expression in CSCC samples (1.00). When analysing the combination of two reference genes for improving stability, NormFinder indicated HPRT1 and ACTB to be the best for MM samples, and HPRT1 and TFRC genes for CSCC. In conclusion, HPRT1 and ACTB genes in combination are the most appropriate choice for normalization in gene expression studies in MM FFPE tissue, while the combination of HPRT1 and TFRC genes are the best option in analysing CSCC FFPE samples. These may be used consistently in forthcoming studies on gene expression in both tumours.
引用
收藏
页码：2167 / 2176
页数：10
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