Parallel genome analysis by two-dimensional DNA typing

Mullaart, E.; Vos, G. J. de; Meerman, Gerard J. te; Uitterlinden, A. G.; Vijg, Jan

doi:10.1038/365469a0

Cited by 22 publications

(12 citation statements)

References 7 publications

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“…Therefore Ingeny (Leiden, The Netherlands) has now developed the IngenoMapper, a 10-gel instrument, in order to better ensure identical running conditions and make handling easier [33].…”

Section: Reproducibilitymentioning

confidence: 99%

Parallel genome analysis by one‐ and two‐dimensional DNA fingerprinting in human gliomas

et al. 1995

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The detection of DNA variation in cancers is an important step in elucidating the mechanism of tumorigenesis. Using the strategy of multipoint genome analysis we detected many differences between glioma-derived and constitutional DNA by customary DNA fingerprinting with simple repetitive oligonucleotide probes. Amplification of the epidermal growth factor receptor (EGFR) gene has been found to be easily detectable as new or highly intensified bands in one-dimensional (1-D) DNA fingerprints of glioblastoma DNA generated with probes (GTG)5 or (GT)8. However, in most low-grade astrocytomas, 1-D DNA fingerprinting has failed to reveal any genomic abnormalities. In these cases a two-dimensional (2-D) technique was successfully employed that is based on size separation in neutral gels followed by sequence-dependent separation in denaturing gradient gels and hybridization with several mini- and microsatellite core probes. The hundreds of spots visualized with this technique were used to detect subtle changes probably occurring as the initial steps of tumorigenesis in human gliomas. On average, five of the approximately 580 spots generated by probes CAC and 33.6 were found to be altered in tumor DNA; 80% of the alterations were spot losses, the rest being spot gains or amplifications. Computer-based image analysis using an external lambda marker provided a stringent way to compare spot patterns generated by 2-D DNA fingerprinting. In comparisons performed between typing patterns generated on the same gel, 99% of truly identical spots were confirmed by the software. In intergel comparisons 84% of identical spots were matched on the basis of the marker information alone.

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“…Therefore Ingeny (Leiden, The Netherlands) has now developed the IngenoMapper, a 10-gel instrument, in order to better ensure identical running conditions and make handling easier [33].…”

Section: Reproducibilitymentioning

confidence: 99%

Parallel genome analysis by one‐ and two‐dimensional DNA fingerprinting in human gliomas

et al. 1995

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“…The apparatus for 2-D DNA typing designed by Mullaart et al (1993) (INGENY B.V.) was used in this study. The PCR product (1 ãl) from each amplified region was mixed with 5 ãl of loading buffer, applied to a neutral 8% polyacrylamide gel (ratio of acrylamide/bis-acrylamide, 37.5:1), and electrophoresed laterally (left to right) at 60°C for 2 hr at 200V in 0.…”

Section: -D Dna Typingmentioning

confidence: 99%

“…However, originally each two-dimensional step had to be done separately, so the procedure was somewhat laborious and low in reproducibility. Mullaart et al (1993) automated a conventional 2-D DNA typing system to avoid manual interference. To decide whether this automated method would be a powerful strategy for mutational analysis, we attempted to establish optimal conditions for screening hMLH1, a human DNA mismatch-repair gene that is constitutionally mutant in many patients with hereditary nonpolyposis colon cancer (HNPCC).…”

Section: Introductionmentioning

confidence: 99%

Mutational analysis of the hMLH1 gene using an automated two-dimensional DNA typing system

et al. 1997

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“…After electrophoresis at 110 V for 17.5 hr at 58"C, the gel was again documented by ethidium bromide staining. Once optimal conditions were established a number of samples were run on an automatic 2D electrophoresis system (Ingeny, Leiden, the Netherlands) (Mullaart et al, 1993), according to the manufacturer's specifications. Conditions were identical with the exception that no manual interference was required to load the first-dimension separation pattern onto the second-dimension gel; switching of directions occurred automatically.…”

Section: Two-dimensional Dna Electrophoresismentioning

confidence: 99%

“…First dimension separation in the upper neutral part of the gel and second dimension separation in a denaturing gradient are carried out consecutively. This was substantially facilitated by using an automated instrument (Mullaart et al, 1993).…”

Section: Detection Formatmentioning

confidence: 99%

Comprehensive and accurate mutation scanning of theCFTR gene by two-dimensional DNA electrophoresis

Hofstra

Scheffer

et al. 1996

Hum. Mutat.

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The large number of possible disease‐causing mutations in the 27 exons of the cystic fibrosis transmembrane conductance regulator (CFTR) gene has severely limited direct diagnosis of cystic fibrosis (CF) patients and carriers by mutation detection. Here we show that in principle testing for mutations in the CFTR gene can be both substantially facilitated and made virtually complete, by two‐dimensional DNA electrophoretic separation of polymerase chain reaction (PCR) amplified exons on the basis of size and basepair sequence in denaturing gradient gels. Under a single optimized set of conditions we were able to obtain a pattern of spots representing all 27 exons of the CFTR gene and to readily detect 17 out of 17 identified sequence variations in 9 different exons in DNA from 11 CF patients and carriers. Our results demonstrate the potential of 2‐dimensional DNA electrophoresis for comprehensive mutation analysis of the CFTR gene. The approach serves as a model for comprehensive diagnosis of the many other large disease genes for which a variety of mutations have also which been reported. © 1996 Wiley‐Liss, Inc.

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Parallel genome analysis by two-dimensional DNA typing

Cited by 22 publications

References 7 publications

Parallel genome analysis by one‐ and two‐dimensional DNA fingerprinting in human gliomas

Parallel genome analysis by one‐ and two‐dimensional DNA fingerprinting in human gliomas

Mutational analysis of the hMLH1 gene using an automated two-dimensional DNA typing system

Comprehensive and accurate mutation scanning of theCFTR gene by two-dimensional DNA electrophoresis

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