Reducing “Double Sequences” in Automated DNA Sequencing with T7 DNA Polymerase and Internal Labeling

Wiemann, Stefan; Schilke, A.; Rechmann, S.; Zimmermann, Juergen; Voß, H.; Ansorge, Wilhelm

doi:10.2144/96205bm12

Cited by 7 publications

(8 citation statements)

References 1 publication

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“…5 and 6). Performing sequencing reactions at high temperatures (cycle sequencing with thermostable polymerases such as Taq, Tth, or SequiTherm, or non-cycling sequencing using IsoTherm) and adding detergents such as Triton X-100 [18], or organic solvents such as DMSO [14,28], can correct such compressions when using dye-primers. The use of dyelabeled dideoxynucleotide terminators also virtually eliminates this problem.…”

Section: Four-color Dna Sequencingmentioning

confidence: 99%

The use of elevated column temperature to extend DNA sequencing read lengths in capillary electrophoresis with replaceable polymer matrices

et al. 1996

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Capillary electrophoresis with a replaceable linear polyacrylamide matrix operated at elevated column temperatures of 55 degrees and 60 degrees C was used to extend the separation of DNA sequencing fragments to lengths greater than 800 bases. A solid-state heater was employed to provide stable, uniform temperature control over a significant portion of the capillary. The polymer matrix, 3% w/v linear polyacrylamide in a denaturing buffer, was replaced in the capillary after each run. Using dye-labeled primers and Sequenase chemistry on an M13mp18 single-stranded template, four-color separations for the sequencing products were obtained, with read lengths in excess of 800 bases. This paper also briefly discusses the effects of buffer denaturants and capillary temperature on separation speed, resolution, and gel compression.

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Section: Four-color Dna Sequencingmentioning

confidence: 99%

The use of elevated column temperature to extend DNA sequencing read lengths in capillary electrophoresis with replaceable polymer matrices

et al. 1996

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“…The detection was chemiluminescent using CSPD (Boehringer) ~G Gz~ Gc~ TAG CaG AGA GGA GGA GGA AGT TTC TGA AGr CAC TGA TAT Z~ GGA for Southern blots, or colorimetric using the NBT/BCIP protocol (B ) [19] hyb GTT ACT GTT GCA ACT GGA TTr CCA AAG TCA CTA TTA ATA GAT TCC TCA GGA AAT CCG GTA USA) following the protocol described in [21]. Electrophoresis and z23 z3z ~4~…”

mentioning

confidence: 99%

Two different respiratory Rieske proteins are expressed in the extreme thermoacidophilic crenarchaeon Sulfolobus acidocaldarius: cloning and sequencing of their genes

1996

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We have isolated two genes encoding Rieske ironRecently we reported the first isolation of a typical Rieske sulfur proteins from the genomic DNA of the thermoacidophilic iron sulfur protein (Rieske-I) from an archaeal source, the crenarchaeon Sulfolobus acidocaldarius (DSM 639). One of the membranes of the extreme thermoacidophilic crenarchaeon genes, named soxL, codes for the previously isolated novel

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“…The use of fluorescein dATP further boosted the read length obtained . The variation in the quality of the sequence noted during routine use (Hawkins et al, 1992) was clarified somewhat after the detailed demands made on the primer and the target sequences were revealed and refined (Wiemann et al, 1996). A screening of the polymerases, which had in the meantime become a target for genetic manipulation (Tabor and Richardson, 1995) showed that by no means all enzymes routinely used in sequencing would accept fluorescent dNTPs as substrates (Voss et al, 1997).…”

Section: B Labelling the Growing Dna Chain (Fig 1b)mentioning

confidence: 99%

Strategies for introducing non-radioactive labels during the automated sequence analysis of nucleic acids

Igloi¹

1998

Electron. J. Biotechnol.

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Rapid, large-scale genome as well as diagnostic DNA sequencing projects are, at present, dependent on the use of sensitive automated sequencers that rely on the detection of fluorescent signals. This emission is not an intrinsic property of the biomolecules but is a property of an optical label that must be incorporated before, during or after the sequence specific reaction. The choice of strategy, that is to be reviewed here, is a function of both the chemistry and the enzymology of the system as well as the nature of the fractionation and the physical parameters of the detection. One may differentiate beween the labelling of the primer, incorporation of the label during elongation or base specific termination with labelled terminators. In reviewing the development of each of these strategies, the conclusion is reached that, whereas labelled primers have universal applicability, the current generation of fluorescent terminators have in, conjunction with appropriate DNA polymerases, attained a refinement that strongly favours their use. However, since they are not available for all commercial sequencing systems, the alternative procedures are not merely of historic interest but are an essential component in DNA sequencing protocols. The possibility of automated direct sequence analysis of RNA has not been ignored completely.

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Reducing “Double Sequences” in Automated DNA Sequencing with T7 DNA Polymerase and Internal Labeling

Abstract: improved method for the isolation of highly polymerized native deoxyribonucleic acid from certain protozoa.

Cited by 7 publications

References 1 publication

The use of elevated column temperature to extend DNA sequencing read lengths in capillary electrophoresis with replaceable polymer matrices

The use of elevated column temperature to extend DNA sequencing read lengths in capillary electrophoresis with replaceable polymer matrices

Two different respiratory Rieske proteins are expressed in the extreme thermoacidophilic crenarchaeon Sulfolobus acidocaldarius: cloning and sequencing of their genes

Strategies for introducing non-radioactive labels during the automated sequence analysis of nucleic acids

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