Reversed-phase high-performance liquid chromatographic system for the rapid, automated purification of oligonucleotides

Birsner, U.; Gilles, U.; Nielsen, P. G.; McMaster, Gary

doi:10.1016/0021-9673(87)80043-2

Cited by 9 publications

(4 citation statements)

References 8 publications

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“…Calf intestine phosphatase was from Boehringer Mannheim. [-y-32P]ATP (>3000 Ci/mmol), [c_-32P]dATP (>3000 Ci/ mmol) and [a-35S]dATP (1200 Ci/mmol) were obtained from Amersham Corp. Oligodeoxynucleotides were synthesized with an automated DNA synthesizer from Applied Biosystems Inc. and HPLC purified using an automated system described previously (Birsner et al, 1987).…”

Section: Methodsmentioning

confidence: 99%

The mouse protein synthesis initiation factor 4A gene family includes two related functional genes which are differentially expressed.

1988

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We have cloned and characterized a family of mouse genomic sequences hybridizing to mouse cDNA probes coding for eIF‐4A, one of the protein synthesis initiation factors involved in the binding of mRNA to the ribosome. We estimate that there is a total of approximately 9‐13 eIF‐4A pseudogenes. We also found an eIF‐4A intronless retroposon which, when compared to the cDNA, contains a single nucleotide difference. This possibly functional gene contains a mouse repetitive B1 element integrated in the promoter region. Furthermore, we have cloned two intron‐containing eIF‐4A genes (termed eIF‐4AI and eIF‐4AII). The eIF‐4AII gene codes for a previously unknown form of eIF‐4A. Northern blot hybridization with RNA from several mouse organs shows a variation in eIF‐4AI expression within a factor of 7. In contrast, relative to liver, eIF‐4AII expression is 20‐ to 30‐times higher in brain and kidney, 10‐ to 17‐fold higher in lung and heart, and is about equally abundant in liver, spleen and thymus. These data suggest that the relative efficiency of protein synthesis initiation for different mRNAs, as reflected by discrimination in messenger 5′‐terminal cap recognition and binding to ribosomes, varies in different tissues.

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Section: Methodsmentioning

confidence: 99%

The mouse protein synthesis initiation factor 4A gene family includes two related functional genes which are differentially expressed.

1988

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“…Oligonucleotides were synthesized with a DNA synthesizer from Applied Biosystems (Foster City, CA) and purified by high-performance liquid chromatography [3]. For the mutagenesis, the recombinant plasmids were transformed into E. coli strain RZ1032 (HfrKL16 PO/45 [lysA(61-62)], dutl, ungl, thil, relA1, Zbd-279::Tn10, supE44), and single-stranded DNA produced in presence of helper phage M13KO7 was used for the mutagenesis [ 14].…”

Section: Site-directed Mutagenesismentioning

confidence: 99%

“…Since the environment around the protein start contained no suitable cloning site, an Nco I site covering the start ATG was created by site-directed mutagenesis with a 21-mer oligonucleotide which exchanged the two bases AG in front of the ATG into CC. The protein coding region was then inserted as Nco I/ I pKK2 3 Hind III fragment into vector pKK233-2. The final construct contained the complete coding region with the deduced start ATG fused to the trc promoter of the vector.…”

Section: Site-directed Mutagenesismentioning

confidence: 99%

Molecular analysis of chalcone and dihydropinosylvin synthase from Scots pine (Pinus sylvestris), and differential regulation of these and related enzyme activities in stressed plants

et al. 1992

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Chalcone synthase (CHS) and stilbene synthase (STS) are closely related polyketide synthases which are key enzymes in the biosynthesis of flavonoids and stilbenes. Scots pine (Pinus sylvestris) is an interesting plant for a direct comparison of the enzymes. It not only contains the usual flavonoids, but also an unusual chalcone derivative (pinocembrin), and it synthesizes stilbenes of the pinosylvin type. We analysed a CHS and a STS by molecular cloning and functional expression in Escherichia coli. The CHS was active not only with 4-coumaroyl-CoA (to naringenin chalcone), but also with cinnamoyl-CoA (leading to pinocembrin). The STS was identified as dihydropinosylvin synthase, because it preferred dihydrocinnamoyl-CoA to cinnamoyl-CoA. The protein deviated in 47 positions from the CHS consensus. It had 73.2% identity with the CHS from P. sylvestris and only 65.3% with a STS from peanut (Arachis hypogaea). We also investigated the regulation of both enzyme types in P. sylvestris plantlets exposed to stress. CHS was present in non-stressed plantlets, and induction led to a transient increase with a peak after 16 h. STS type activities were regulated differently and were absent in non-stressed plantlets. Increases were observed after a lag period of at least 6 h, and highest activities were obtained after 30 h. The analysis of the reactions in the plant extracts and the substrate specificity of the cloned STS indicate that the plants contain at least two different types of STS: the cloned dihydropinosylvin synthase and a pinosylvin synthase which preferentially utilizes cinnamoyl-CoA as substrate.

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“…Mass., USA) [17], Labeling with [y-! :P|-ATP was done in a kinase reaction, as described by Schafer et al [15].…”

Section: Methodsmentioning

confidence: 99%

New Mutation versus Exclusion at the Alpha-1-Antitrypsin Locus: A Multif aceted Approach in a Problematical Paternity Case

Bender¹,

Kasulke

Mayerová³

et al. 1991

Hum Hered

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In a case of disputed paternity with overwhelming indications of fatherhood for the putative father, as supported by serological tests and biostatistical evaluation, a classical exclusion constellation was found at the α₁-antitrypsin (PI) locus: mother PI M1; child PI M1M3, and putative father PI M1M2. Additional studies included PI oligonucleotide phenotyping and DNA fingerprint analysis. Results from the entire data set led us to assume a rare genetic event at the paternal PI locus. Intracistronal crossing-over offered the most parsimonious explanation, and was compatible with the PI gene DNA sequence and the amino acid sequences of the molecule and its allelic forms, as well as with the experimental findings.

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Reversed-phase high-performance liquid chromatographic system for the rapid, automated purification of oligonucleotides

Cited by 9 publications

References 8 publications

The mouse protein synthesis initiation factor 4A gene family includes two related functional genes which are differentially expressed.

The mouse protein synthesis initiation factor 4A gene family includes two related functional genes which are differentially expressed.

Molecular analysis of chalcone and dihydropinosylvin synthase from Scots pine (Pinus sylvestris), and differential regulation of these and related enzyme activities in stressed plants

New Mutation versus Exclusion at the Alpha-1-Antitrypsin Locus: A Multif aceted Approach in a Problematical Paternity Case

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