cDNA cloning is a central technology in molecular biology. cDNA sequences are used to determine mRNA transcript structures, including splice junctions, open reading frames (ORFs) and 5'- and 3'-untranslated regions (UTRs). cDNA clones are valuable reagents for functional studies of genes and proteins. Expressed Sequence Tag (EST) sequencing is the method of choice for recovering cDNAs represent...

Choline acetyltransferase (EC 2.3.1.6) is the biosynthetic enzyme for the neurotransmitter acetylcholine. To isolate choline acetyltransferase cDNA clones, a cDNA library was constructed from poly(A)+ RNA of Drosophila melanogaster heads, these being one of the richest known sources of the enzyme. By screening the cDNA library with a mixture of three different monoclonal antibodies to Drosophil...

We have used a biotin-based subtraction procedure to enrich a potato tuber cDNA library for sink-specific clones. The method uses single-stranded phagemids with directional inserts as both driver and target. We modified the XZAP II vector for the directional cDNA cloning and for subsequent subtractive hybridization. This improved method of subtractive hybridization circumvents many problems fac...

cDNA cloning very often involves screening of a library by hybridization with a short fragment of the desired cDNA as a probe. Commonly the choice of the library is made on the basis of the properties of the vector, the extent of expression in the tissue from which the library is made, the ‘quality ’ of the library, and possibly other parameters. However, even when these choices are optimally m...

Aim Kampo medicine is composed of a variety crude drugs. Each drug has its own effects, and the interaction all drugs produces therapeutic effect medicine. In order to examine characteristics drugs, 120 types herbal medicines were characterized based on their three-dimensional high-performance liquid chromatography (3D-HPLC) profiles as well effects cell morphology/proliferation when added cult...

cDNA species coding for α-methylacyl-CoA racemase were cloned from rat and mouse liver cDNA libraries and characterized. The rat liver λgt11 cDNA expression library was screened with anti-racemase IgG [Schmitz, Albers, Fingerhut and Conzelmann (1995) Eur. J. Biochem. 231, 815–822]. Several fulllength clones were obtained that contained an open reading frame of 1083 bp, coding for a protein of 3...

Monospecific antibodies directed against the chicken progesterone receptor (PR) form B were used to screen a randomly primed phage lambda gt11 cDNA expression library prepared from size-fractionated chicken oviduct mRNA. Two independent immunoreactive clones, lambda cPR1 and lambda cPR2, were isolated. Antibodies selected from anti-PR form B antiserum on matrices of lambda cPR1 and lambda cPR2 ...

In a cellular transcriptome, the number of mRNA copies per gene may differ by several orders. In cDNA libraries, performed from mRNA, these proportions are the same. Normalization methods allow us to equalize numbers of gene’s copies in the library. Normalized cDNA libraries are used to discover new genes transcribed at relatively low levels or for functional screenings. Here, we observed diffe...

A molecular map has been constructed for the rice genome comprised of 726 markers (mainly restriction fragment length polymorphisms; RFLPs). The mapping population was derived from a backcross between cultivated rice, Oryza sativa, and its wild African relative, Oryza longistaminata. The very high level of polymorphism between these species, combined with the use of polymerase chain reaction-am...