Distinguishing between mouse IL-3 and IL-3 receptor-like (IL-5/GM-CSF receptor converter) mRNAs using the polymerase chain reaction method.
A set of primers (MF43, MF44 and MF45) were designed and used in the polymerase chain reaction to distinguish between the expression of mouse IL-3 receptor and mouse IL-3 receptor-like mRNAs. Primers MF43 and MF45 were specific for IL-3 receptor mRNA while the primers MF44 and MF45 were specific for IL-3 receptor-like mRNA. Primers MF44 and MF45 could not amplify IL-3 receptor cDNA even at an annealing temperature of 46 degrees C which is 20 degrees C below the melting temperature of the primers, or at high template concentrations (up to 100 ng cDNA). The optimal range of Mg2+ concentrations for the two pairs of primers MF43, MF45 and MF44, MF45, were essentially the same and this permits comparisons of the expression level of these two mRNAs under identical PCR conditions. Both the IL-3 receptor and IL-3 receptor-like mRNAs could be detected in normal bone marrow cells and IL-3-dependent cell lines (FDC-P1 and 32D cl-23), as well as in the IL-3 independent cell lines P388D1 and WEHI-3B, the latter being a constitutive producer of IL-3. In contrast, neither species of mRNA was detected in the T lymphoma cell line (EL-4). The ratio of IL-3 receptor-like mRNA to IL-3 receptor mRNA was usually greater than 1, except in 32D cl-23 cells where it was 0.66.
Fung
M C
MC
Division of Biochemistry and Molecular Biology, John Curtin School of Medical Research, Australian National University, Canberra City.
Mak
N K
NK
Leung
K N
KN
Hapel
A J
AJ
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D015342:DNA Probes; D004305:Dose-Response Relationship, Drug; D015870:Gene Expression; D008274:Magnesium; D051379:Mice; D008811:Mice, Inbred DBA; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D012333:RNA, Messenger / Q000096:biosynthesis; D011971:Receptors, Immunologic / Q000096:biosynthesis*; D018123:Receptors, Interleukin; D016185:Receptors, Interleukin-3 / Q000096:biosynthesis*
D015342:DNA Probes; D012333:RNA, Messenger; D011971:Receptors, Immunologic; D018123:Receptors, Interleukin; D016185:Receptors, Interleukin-3; C074452:interleukin 3 receptor-like molecule; D008274:Magnesium