Evidence that TSG101 aberrant transcripts are PCR artifacts.
Critical analysis of the data published so far concerning the TSG101 gene revealed some inconsistencies leading us to its re-evaluation in 80 breast, brain, colon, and neuroectodermal tumors and 37 normal tissue specimens. In this study, the occurrence of TSG101 cDNA aberrant transcripts was verified, but in addition we made observations that are in apparent conflict with the aberrant splicing theory supposed as the underlying mechanism for transcript formation: the location of most deletion breakpoints within exons and nonconformity of these putative splice sites with the highly conserved GT-AG rule, detection of insertions as well as nonreproducible and highly variable results in repeated RT-nested PCRs. Furthermore, we found that reamplification of full-length TSG101 cDNA products leads to the generation of deleted transcripts. In summary, for the first time we provide evidence that the acquired TSG101 transcripts are caused by PCR artifacts.
Hampl
M
M
Department of General/Thoracic and Vascular Surgery, Technical University of Dresden, Germany.
Hampl
J
J
Plaschke
J
J
Fitze
G
G
Schackert
G
G
Saeger
H D
HD
Schackert
H K
HK
D016477:Artifacts; D001482:Base Composition; D001483:Base Sequence; D015139:Blotting, Southern; D001932:Brain Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D003110:Colonic Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D004268:DNA-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics*; D056827:Endosomal Sorting Complexes Required for Transport; D005260:Female; D006801:Humans; D016350:Leucine Zippers; D008969:Molecular Sequence Data; D009361:Neoplasm Invasiveness; D009362:Neoplasm Metastasis; D009447:Neuroblastoma / Q000235:genetics / Q000378:metabolism; D017599:Neuroectodermal Tumors / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D016133:Polymerase Chain Reaction / Q000379:methods*; D012016:Reference Values; D017384:Sequence Deletion; D014157:Transcription Factors / Q000096:biosynthesis* / Q000235:genetics*; D014158:Transcription, Genetic; D000099177:Tsg101 Protein
D004268:DNA-Binding Proteins; D056827:Endosomal Sorting Complexes Required for Transport; D014157:Transcription Factors; D000099177:Tsg101 Protein