Optical Mapping of BAC Clones from the Human Y Chromosome <i>DAZ</i> Locus

Giacalone, Joseph; Delobette, Stephanie; Gibaja, Veronica; Ni, Lei; Skiadas, Yiannis; Qi, Rong; Edington, Joanne; Lai, Zhongwu; Gebauer, Damara; Zhao, Hongjuan; Anantharaman, Thomas; Mishra, Bhubaneswar; Brown, Laura G.; Saxena, Richa; Page, David C.; Schwartz, David C.

doi:10.1101/gr.112100

Cited by 15 publications

(12 citation statements)

References 32 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…However, new technologies are being developed to provide much greater resolution (Ͻ50 kb), including "genome tiling" BAC arrays, 77 digital karyotyping, 78 and optical mapping. 79 These new techniques may well open up a previously inaccessible avenue of investigation into the genomic basis of SCD, and complex phenotypes in general.…”

Section: Arking Et Al Genomics In Sudden Cardiac Death 719mentioning

confidence: 99%

Genomics in Sudden Cardiac Death

Arking

Chugh

Chakravarti

et al. 2004

Circulation Research

View full text Add to dashboard Cite

Abstract-Sudden cardiac death (SCD) remains a public health problem of major magnitude. Contrary to earlier expectations, and despite decreased overall cardiac mortality, SCD rates appear to be rising in concert with escalating global prevalence of coronary disease and heart failure, the two major conditions predisposing to SCD. With the exception of the implantable defibrillator, there are few effective approaches to SCD prevention and even fewer clues concerning patient phenotypes predisposed to life-threatening arrhythmias. Clinical variables such as ejection fraction predict mortality but are not sensitive enough to identify many high SCD risk patients. The predictive power of autonomic dysregulation and markers such as lipid levels, hypertension, diabetes, and smoking is quite low in subclinical heart disease, the population in which the majority of SCDs occur. This review addresses advances in genomic science applicable to the SCD public health problem in both rare and common forms of heart disease. These include novel bioinformatic approaches to both identify candidate genes/pathways and identify previously unknown functional genetic elements, as well as methods to comprehensively screen these elements. We also discuss the possibility of applying high-density genome-wide SNP analyses to examine genetic contributions to arrhythmia susceptibility in community-based, case-control studies of common forms of SCD. The development of novel strategies to identify contributors to susceptibility in common cardiac phenotypes is most likely to lead to new and relevant therapeutic targets for SCD.

show abstract

Section: Arking Et Al Genomics In Sudden Cardiac Death 719mentioning

confidence: 99%

Genomics in Sudden Cardiac Death

Arking

Chugh

Chakravarti

et al. 2004

Circulation Research

View full text Add to dashboard Cite

show abstract

“…While the Optical Mapping system has been developed into a robust, high throughput, single molecule system able to construct physical maps of whole genomes, including microbial, fungal, plant and mammalian genomes [18,19,34,[44][45][46][47][48][49], it has now been extended to analyze the positions and fluorescence intensities of in vitro transcription products associated with surface-bound single template DNA molecules. This new method allows rapid analysis of important transcription caveats on OM surfaces.…”

Section: Discussionmentioning

confidence: 99%

Transchip: Single-molecule detection of transcriptional elongation complexes

Schwartz

2007

Analytical Biochemistry

Self Cite

View full text Add to dashboard Cite

A new single molecule system -Transchip -was developed for analysis of transcription products at their genomic origins. The bacteriophage T7 RNA polymerase and its promoters were used in a model system, and resultant RNAs were imaged and detected at their positions along single template DNA molecules. This system, Transchip, has drawn from critical aspects of Optical Mapping, a single molecule system that enables the construction of high resolution, ordered restriction maps of whole genomes from single DNA molecules. Through statistical analysis of hundreds of single molecule template/transcript complexes, Transchip enables analysis of the locations and strength of promoters, the direction and processivity of transcription reactions and termination of transcription. These novel results suggest that the new system may serve as a high-throughput platform to investigate transcriptional events on a large, genome-wide scale. KeywordsSingle molecule; transcription; elongation complex; T7 RNA polymerase; promoter mapping; automated fluorescence microscopy Introductory StatementThis study aims to develop an integrated system for transcriptional analysis of single template DNAs, with the ultimate goal being analysis of transcriptional regulation for genes on a whole genome basis. Large template DNA molecules can be queried to probe for regulatory elements and for transcriptional events. The Transchip system described in this manuscript aims to produce large and meaningful datasets for transcriptional analysis, all based around single molecule measurements. Experiments based on single molecules are direct and rapid, and can generate a read-out of thousands of data points in a short time period.The development of single molecule approaches to biochemical analysis has enabled a broad range of techniques that visualize the details of and that elegantly probe individual steps involved in transcription [1][2][3][4][5][6]. These elegant biochemical studies performed on single molecules have brought numerous insights into the mechanisms of transcription, but they do not lend themselves to high throughout transcriptional analysis. Visualization and mapping of RNAs transcribed from single template molecules has long been possible by electron + To whom correspondence should be addressed: dcschwartz@wisc.edu, 608-265-0546; 608-265-6743 (FAX) # current position, Amgen, Inc., One

show abstract

“…As the optical mapping became a tool for construction of restriction mega-maps (Giacalone et al 2000), it also became very desirable to have a very rare and reliable restriction site on the vector for efficient and convenient linearization of clones. To meet this goal, we used the very rare restriction site, I-SceI, for the intron homing endonuclease (Monteilhet et al 1990) and constructed pBAC/oriV/SceI by cloning the NotIless oriV fragment (see legend to Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Further improvements include the following: (1) Incorporation of the I-SceI site into pBAC/oriV derivatives creates clones more suitable for the optical mapping (Giacalone et al 2000). Our vectors pBAC/oriV/SceI and pTrueBlue-BAC2/ oriV/SceI were designed for that purpose.…”

Section: Discussionmentioning

confidence: 99%

Conditionally Amplifiable BACs: Switching From Single-Copy to High-Copy Vectors and Genomic Clones

Wild¹,

Hradečná²,

Szybalski³

2002

Genome Res.

167

145

View full text Add to dashboard Cite

The widely used, very-low-copy BAC (bacterial artificial chromosome) vectors are the mainstay of present genomic research. The principal advantage of BACs is the high stability of inserted clones, but an important disadvantage is the low yield of DNA, both for vectors alone and when carrying genomic inserts. We describe here a novel class of single-copy/high-copy (SC/HC) pBAC/oriV vectors that retain all the advantages of low-copy BAC vectors, but are endowed with a conditional and tightly controlled oriV/TrfA amplification system that allows: (1) a yield of ∼ 100 copies of the vector per host cell when conditionally induced with L-arabinose, and (2) analogous DNA amplification (only upon induction and with copy number depending on the insert size) of pBAC/oriV clones carrying >100-kb inserts. Amplifiable clones and libraries facilitate high-throughput DNA sequencing and other applications requiring HC plasmid DNA. To turn on DNA amplification, which is driven by the oriV origin of replication, we used copy-up mutations in the gene trfA whose expression was very tightly controlled by the araC-P araBAD promoter/regulator system. This system is inducible by L-arabinose, and could be further regulated by glucose and fucose. Amplification of DNA upon induction with L-arabinose and its modulation by glucose are robust and reliable. Furthermore, we discovered that addition of 0.2% D-glucose to the growth medium helped toward the objective of obtaining a real SC state for all BAC systems, thus enhancing the stability of their maintenance, which became equivalent to cloning into the host chromosome.

show abstract

Optical Mapping of BAC Clones from the Human Y Chromosome DAZ Locus

Cited by 15 publications

References 32 publications

Genomics in Sudden Cardiac Death

Genomics in Sudden Cardiac Death

Transchip: Single-molecule detection of transcriptional elongation complexes

Conditionally Amplifiable BACs: Switching From Single-Copy to High-Copy Vectors and Genomic Clones

Contact Info

Product

Resources

About