Detection of differentially expressed gelatinase A in metastatic and non-metastatic subpopulations of tumor cells by target RNA arbitrarily primed polymerase chain reaction (TRAP-PCR)
- Resource Type
- Authors
- Laia Masramon; Angels Fabra; Miquel Angel Peinado; Marc Adrover; Antonia Vinyals; Milagro Gonzalez-Garrigues; Ana Llorens; Pedro Alía
- Source
- Clinicalexperimental metastasis. 16(7)
- Subject
- Cancer Research
Gelatinase A
Gelatin Zymography
Biology
Polymerase Chain Reaction
Sensitivity and Specificity
law.invention
Metastasis
Mice
law
Complementary DNA
medicine
Tumor Cells, Cultured
Non metastatic
Animals
Neoplasm Invasiveness
Northern blot
RNA, Messenger
Polymerase chain reaction
RNA
Mammary Neoplasms, Experimental
Metalloendopeptidases
General Medicine
medicine.disease
Blotting, Northern
Molecular biology
Oncology
Gelatinases
Matrix Metalloproteinase 2
Female
- Language
- ISSN
- 0262-0898
We have developed a novel procedure called Targeted RNA AP-PCR (TRAP-PCR) to quantitatively measure specific mRNA expression. The target mRNA is reverse transcribed using a specific primer and PCR is performed under low stringency conditions to generate a rich fingerprint-type band pattern. In this situation multiple sequences are coamplified with the targeted sequence. The amplification is carried out in a competitive fashion and is, in consequence, quantitative. We have applied this technique to determine Gelatinase A (Gel A) mRNA expression in the MXT mouse mammary carcinoma system. TRAP-PCR analysis using primers for Gel A produced a reproducible fingerprint including one major band whose iden-tity was confirmed to be Gel A cDNA. Highly metastatic MXT subclones show an increased Gel A expres-sion. Results were confirmed by Northern blot and protein activity (gelatin zymography). TRAP-PCR is a simple, sensitive and specific technique to comparatively quantify mRNA expression and requires less template than conventional methods. ©Lippincott Williams & Wilkins