Software for Automated Analysis of DNA Fingerprinting Gels

Fuhrmann, D.R.; Krzywinski, Martin; Chiu, Readman; Saeedi, Parvaneh; Schein, Jacqueline E.; Bosdet, Ian; Chinwalla, Asif T.; Hillier, La Deana W.; Waterston, Robert H.; Mcpherson, John Douglas; Jones, Steven J.M.; Marra, Marco A.

doi:10.1101/gr.904303

Cited by 35 publications

(28 citation statements)

References 24 publications

(26 reference statements)

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“…Gel images were acquired using a Molecular Dynamics Fluorimager 595. Lane tracking of the digitized gel images was manually adjusted using Image software (www.sanger.ac.uk/resources/software/image/), and restriction fragments were identified and sized automatically using BandLeader software (Fuhrmann et al 2003).…”

Section: Bac Restriction Fingerprintingmentioning

confidence: 99%

The Release 6 reference sequence of the Drosophila melanogaster genome

et al. 2015

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Drosophila melanogaster plays an important role in molecular, genetic, and genomic studies of heredity, development, metabolism, behavior, and human disease. The initial reference genome sequence reported more than a decade ago had a profound impact on progress in Drosophila research, and improving the accuracy and completeness of this sequence continues to be important to further progress. We previously described improvement of the 117-Mb sequence in the euchromatic portion of the genome and 21 Mb in the heterochromatic portion, using a whole-genome shotgun assembly, BAC physical mapping, and clone-based finishing. Here, we report an improved reference sequence of the single-copy and middle-repetitive regions of the genome, produced using cytogenetic mapping to mitotic and polytene chromosomes, clone-based finishing and BAC fingerprint verification, ordering of scaffolds by alignment to cDNA sequences, incorporation of other map and sequence data, and validation by whole-genome optical restriction mapping. These data substantially improve the accuracy and completeness of the reference sequence and the order and orientation of sequence scaffolds into chromosome arm assemblies. Representation of the Y chromosome and other heterochromatic regions is particularly improved. The new 143.9-Mb reference sequence, designated Release 6, effectively exhausts clone-based technologies for mapping and sequencing. Highly repeat-rich regions, including large satellite blocks and functional elements such as the ribosomal RNA genes and the centromeres, are largely inaccessible to current sequencing and assembly methods and remain poorly represented. Further significant improvements will require sequencing technologies that do not depend on molecular cloning and that produce very long reads.

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Section: Bac Restriction Fingerprintingmentioning

confidence: 99%

The Release 6 reference sequence of the Drosophila melanogaster genome

et al. 2015

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“…Fingerprints were generated by digesting RHA1 fosmid clones with BamHI and separating the resulting fragments on 1.2% agarose gels (40, 52a). Gel images were processed and captured with IMAGE (http://www-.sanger.ac.uk/Software/Image), and the fragments were called using BAN-DLEADER software (22). A total of 4,973 fingerprints were obtained from 4992 fosmid clones that were analyzed.…”

Section: Chemicalsmentioning

confidence: 99%

Functional Characterization of a Catabolic Plasmid from Polychlorinated- Biphenyl-Degrading Rhodococcus sp. Strain RHA1

Warren¹,

Hsiao

Kudo

et al. 2004

J Bacteriol

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Rhodococcus sp. strain RHA1, a potent polychlorinated-biphenyl (PCB)-degrading strain, contains three linear plasmids ranging in size from 330 to 1,100 kb. As part of a genome sequencing project, we report here the complete sequence and characterization of the smallest and least-well-characterized of the RHA1 plasmids, pRHL3. The plasmid is an actinomycete invertron, containing large terminal inverted repeats with a tightly associated protein and a predicted open reading frame (ORF) that is similar to that of a mycobacterial rep gene. The pRHL3 plasmid has 300 putative genes, almost 21% of which are predicted to have a catabolic function. Most of these are organized into three clusters. One of the catabolic clusters was predicted to include limonene degradation genes. Consistent with this prediction, RHA1 grew on limonene, carveol, or carvone as the sole carbon source. The plasmid carries three cytochrome P450-encoding (CYP) genes, a finding consistent with the high number of CYP genes found in other actinomycetes. Two of the CYP genes appear to belong to novel families; the third belongs to CYP family 116 but appears to belong to a novel class based on the predicted domain structure of its reductase. Analyses indicate that pRHL3 also contains four putative "genomic islands" (likely to have been acquired by horizontal transfer), insertion sequence elements, 19 transposase genes, and a duplication that spans two ORFs. One of the genomic islands appears to encode resistance to heavy metals. The plasmid does not appear to contain any housekeeping genes. However, each of the three catabolic clusters contains related genes that appear to be involved in glucose metabolism.Rhodococcus is a widely occurring genus of aerobic, nonmotile soil bacteria that are closely related to three other genera of GC-rich actinomycetes: Gordonia, Nocardia, and Mycobacterium. Rhodococci degrade an extraordinarily wide variety of organic substrates and thus play an important role in the global C cycle. The unusual armamentarium of enzymatic activities involved in these processes has been exploited in applications ranging from commodity chemical production to the desulfurization of fossil fuels (6). Consequently, the metabolic capabilities of rhodococci are of interest to the pharmaceutical, environmental, chemical, and energy sectors.Rhodococcus sp. strain RHA1 is characterized by its exceptional ability to transform polychlorinated biphenyls (PCBs) (53), a particularly widespread and persistent class of environmental pollutants. It is generally thought that in aerobic bacteria, PCBs are cometabolized by the bph pathway, which is responsible for the aerobic degradation of biphenyl (23). The upper bph pathway consists of four enzymatic activities that together transform biphenyl to benzoate and 2-hydroxypenta-2,4-dienoate. For each of these four steps, RHA1 appears to possess multiple isozymes, which may help explain the strain's superior PCB-transforming capabilities. Thus, the strain contains at least three bph-type ring-hydroxylating di...

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“…Even with assistance from Image software (Sulston et al, 1989), this is a time-consuming process, which also requires considerable skill; however, an automated alternative has recently been described by Fuhrmann et al (2003). Another drawback of agarose fingerprinting is that the fingerprints contain relatively little information; this is because the fragments are large and few in number and their sizing is not precise.…”

mentioning

confidence: 99%

Whole-Genome Validation of High-Information-Content Fingerprinting

Bharti²,

et al. 2005

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Fluorescent-based high-information-content fingerprinting (HICF) techniques have recently been developed for physical mapping. These techniques make use of automated capillary DNA sequencing instruments to enable both high-resolution and high-throughput fingerprinting. In this article, we report the construction of a whole-genome HICF FPC map for maize (Zea mays subsp. mays cv B73), using a variant of HICF in which a type IIS restriction enzyme is used to generate the fluorescently labeled fragments. The HICF maize map was constructed from the same three maize bacterial artificial chromosome libraries as previously used for the whole-genome agarose FPC map, providing a unique opportunity for direct comparison of the agarose and HICF methods; as a result, it was found that HICF has substantially greater sensitivity in forming contigs. An improved assembly procedure is also described that uses automatic end-merging of contigs to reduce the effects of contamination and repetitive bands. Several new features in FPC v7.2 are presented, including shared-memory multiprocessing, which allows dramatically faster assemblies, and automatic end-merging, which permits more accurate assemblies. It is further shown that sequenced clones may be digested in silico and located accurately on the HICF assembly, despite size deviations that prevent the precise prediction of experimental fingerprints. Finally, repetitive bands are isolated, and their effect on the assembly is studied.

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Software for Automated Analysis of DNA Fingerprinting Gels

Cited by 35 publications

References 24 publications

The Release 6 reference sequence of the Drosophila melanogaster genome

The Release 6 reference sequence of the Drosophila melanogaster genome

Functional Characterization of a Catabolic Plasmid from Polychlorinated- Biphenyl-Degrading Rhodococcus sp. Strain RHA1

Whole-Genome Validation of High-Information-Content Fingerprinting

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