title: Template switching within exons 3 and 4 of KV11.1 (HERG) gives rise to a 5' truncated cDNA.
abstract: K(V)11.1 (HERG) channels contribute to membrane potential in a number of excitable cell types. We cloned a variant of K(V)11.1 from human jejunum containing a 171 bp deletion spanning exons 3 and 4. Expression of a full-length cDNA clone containing this deletion gave rise to protein that trafficked to the cell membrane and generated robust currents. The deletion occurred in a G/C-rich region and identical sequence elements of UGGUGG were located at the deletion boundaries. In recent studies these features have been implicated to cause deletions via template switching during cDNA synthesis. To examine this possibility we compared cDNAs from human brain, heart, and jejunum synthesized at lower (42 degrees C) and higher temperatures (70 degrees C). The 171 bp deletion was absent at the higher temperature. Our results suggest that the sequence and secondary structure of mRNA in the G/C rich region leads to template switching producing a cDNA product with a 171 bp deletion.
authors.affiliation: Department of Physiology and Cell Biology, University of Nevada School of Medicine, Reno, 89557, USA.
authors.forename: S
authors.initials: S
authors.lastname: Ro
authors.forename: S H
authors.initials: SH
authors.lastname: Kang
authors.forename: A M
authors.initials: AM
authors.lastname: Farrelly
authors.forename: T
authors.initials: T
authors.lastname: Ordog
authors.forename: R
authors.initials: R
authors.lastname: Partain
authors.forename: N
authors.initials: N
authors.lastname: Fleming
authors.forename: K M
authors.initials: KM
authors.lastname: Sanders
authors.forename: J L
authors.initials: JL
authors.lastname: Kenyon
authors.forename: K D
authors.initials: KD
authors.lastname: Keef
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D001921:Brain / Q000378:metabolism; D002460:Cell Line; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D005091:Exons / Q000235:genetics*; D020869:Gene Expression Profiling; D006639:Histidine / Q000235:genetics / Q000276:immunology; D006801:Humans; D007583:Jejunum / Q000378:metabolism; D008564:Membrane Potentials / Q000235:genetics / Q000502:physiology; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D009206:Myocardium / Q000378:metabolism; D009690:Nucleic Acid Conformation; D024642:Potassium Channels, Voltage-Gated / Q000235:genetics*; D012313:RNA / Q000737:chemistry / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017384:Sequence Deletion; D013696:Temperature; D013698:Templates, Genetic; D014158:Transcription, Genetic / Q000235:genetics*; D014162:Transfection
chemical_list: D018076:DNA, Complementary; C489610:KCNV2 protein, human; D024642:Potassium Channels, Voltage-Gated; D006639:Histidine; D012313:RNA