Time-dependent RNA degradation affecting cDNA array quality in spontaneous canine tumours sampled using standard surgical procedures.
Heterogeneous gene expression in tumours and the degradation of RNA when sampling under non-RNAse-free conditions may limit the potential benefit of cDNA array studies. This study examines changes in the integrity of RNA by means of RNA gel electrophoresis at various post-operative intervals on canine mammary tumours (n=10) and malignant lymphoma (n=1). The tumours were cut into pieces (3-5 mm diameter, approximately 50 mg) and kept in tubes without RNAse-free buffer at room temperature. No special precautions were taken to avoid the influences of Rnase; rather, normal surgical procedures were used. We found that total RNA of the mammary tumours started to degrade within 30 min of the operation, and the rate of degradation increased up to 4 h, which was the last time point included in this study. RNA in the lymphoma tumours degraded more rapidly, and was completely degraded at 30 min post-operation. The degradation of mRNA in the mammary tumours, as studied by human cDNA arrays, was heterogeneous, i.e. some mRNA degraded completely, some only partially. This indicates that the mRNA degradation rate varied depending on the type of mRNA. However, since we found that gene expression differs depending on the part of the mammary tumour examined, one cannot exclude that the variation in the mRNA degradation rate may simply reflect heterogeneous gene expression within the tumour. We conclude that RNA integrity is unaffected immediately after sampling under non-RNAse-free conditions; however, the tumour sample should be preserved under RNAse-free conditions within 15 min to avoid RNA degradation. This is a much shorter time interval than previously reported in other similar studies; however, these studies generally treated normal tissue, under which 3-5 h non-RNAse-free conditions have been found not to affect RNA quality.
Von Euler
Henrik
H
Department of Small Animal Clinical Sciences, Swedish University of Agricultural Sciences (SLU), Uppsala, Sweden.
Khoshnoud
Reza
R
He
Qimin
Q
Khoshnoud
Aida
A
Fornander
Tommy
T
Rutqvist
Lars-Erik
LE
Skog
Sven
S
D000818:Animals; D001943:Breast Neoplasms / Q000235:genetics / Q000473:pathology / Q000601:surgery / Q000662:veterinary; D004283:Dog Diseases / Q000235:genetics* / Q000473:pathology / Q000601:surgery; D004285:Dogs; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D016393:Lymphoma, B-Cell; D009369:Neoplasms / Q000235:genetics* / Q000473:pathology / Q000601:surgery / Q000662:veterinary*; D020411:Oligonucleotide Array Sequence Analysis / Q000592:standards*; D020871:RNA Stability; D012333:RNA, Messenger / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D013997:Time Factors
D012333:RNA, Messenger