Isolation and characterisation of the human lung NK-2 receptor gene using rapid amplification of cDNA ends.
Functional cDNA clones for human NK-2 receptor were isolated from human lung RNA using a polymerase chain reaction (PCR) based method (RACE-PCR). In this method the cDNA was isolated as 5' end and 3'-end fragments; the entire cDNA was obtained by RNA-PCR. The sequence derived was 398 amino acids in length encoding an open-reading frame that was highly homologous to both the bovine and rat NK-2 receptor. The entire human cDNA sequence was cloned into a mammalian expression vector and mRNA was synthesised by in vitro transcription. Applications of tachykinins caused membrane current responses in Xenopus oocytes injected with the in vitro synthesised mRNA. The most potent of the three tachykinin peptides tested was neurokinin A. We have screened a human cosmid library and isolated a clone which contains the entire NK-2 receptor gene. The gene contains five exons and we have determined the complete sequence of the exons and the intron-exon junctions.
Graham
A
A
Biotechnology Department, ICI Pharmaceuticals, Macclesfield, Cheshire, UK.
Hopkins
B
B
Powell
S J
SJ
Danks
P
P
Briggs
I
I
D000368:Aged; D000595:Amino Acid Sequence; D001483:Base Sequence; D004247:DNA / Q000235:genetics* / Q000302:isolation & purification; D005796:Genes; D006801:Humans; D008168:Lung / Q000502:physiology*; D008297:Male; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D016133:Polymerase Chain Reaction / Q000379:methods; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D012333:RNA, Messenger / Q000235:genetics; D017981:Receptors, Neurotransmitter / Q000235:genetics*; D018039:Receptors, Tachykinin; D015183:Restriction Mapping; D015320:Tachykinins / Q000378:metabolism
D015345:Oligonucleotide Probes; D012333:RNA, Messenger; D017981:Receptors, Neurotransmitter; D018039:Receptors, Tachykinin; D015320:Tachykinins; D012313:RNA; D004247:DNA