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Test with OpGen technology

Test with OpGen technology.

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Test with OpGen technology

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  1. Test with OpGentechnology “Optical Mapping is a de novo process that generates whole genome, ordered, restriction maps with no requirement for previous sequence information. Whole genome analysis may be performed on bacteria, yeast or fungi. Where current technologies like microarray or SNP genotyping systems detect small changes in DNA sequence, Optical Mapping can find the unexpected — genomic rearrangements and modifications to genomic architecture and novel discovery, independent of prior sequence information.” Observed for Ae. tauschiiBAC clones Expected

  2. “Nano-Mapping” BioNanomatrix: “The core technology is a disposable nanochannel array "chip" that, in conjunction with other system components, permits fast and efficient processing, labeling, detection, and analysis of very long segments of DNA and other biomolecules.”

  3. Nick-Labeling • Nicking endonuclease recognizes DNA sequence motif and cuts only one of the DNA strands • Nb.BbvC1 nicks DNA at GCTCAGG • Fluorescent dye-terminator incorporated at nicking site (Nt.BbvCI is a nicking endonuclease that cleaves only one strand of DNA on a double-stranded DNA substrate.)

  4. GCTCAGG Motif Map of Lambda 12.7 kb 4 kb 13 kb 10.5 kb 8 kb ++ +++

  5. GCTCAGG Motif Map of Lambda 42.1kbp 18.2kbp 12.6kbp 32.2kbp 41398bp 30519bp 17244bp 12476bp Nucleic Acids Res. 2007; 35:e16

  6. Nick-Flap Labeling

  7. DNA Linearization and Imaging

  8. Data collection About 1 Gb raw data from one device in 5 scans (20 minutes)

  9. De novo Clustering

  10. Clusters in Opposite Orientation 150kb 150kb

  11. Histogram Plot Consensus Map Reference Human genome data

  12. Cluster Mapped to human Chr 7 170kb

  13. De Novo Sequence Assembly

  14. 3048-Cre linearization (90% efficiency) • 50nM lox/ 1 nM BAC • 1h 37C • 100nM lox/ 1 nM BAC • 200nM lox/ 1 nM BAC kb Test with Ae.tauschii BAC clones circular linearized

  15. Individual clone maps from clone mixture Test with Ae.tauschii BAC clones

  16. Cone D1 HI064J24 BbvCI BspQI Start of vector Linearized by loxp 2kb extra Asisi site BspQI BbvCI 2kb extra Linearized by AsiSi

  17. BSPQI GCTCTTC BbvcI GCTGAGG Reference (bp) Nano map Reference (bp) Nano map 1.3kb extra Sequences? 2kb extra Sequences? The total length detected us is 108540bp Your reference is 98800, total is 10600bp(+8000bp of vector) We detected 2kb extra sequences, which Should be in the above region

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