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Test with opgen technology

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


Nano mapping

“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.”


Nick labeling
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.)


Test with opgen technology

GCTCAGG Motif Map of Lambda

12.7 kb

4 kb

13 kb

10.5 kb

8 kb

++

+++


Test with opgen technology

GCTCAGG Motif Map of Lambda

42.1kbp

18.2kbp

12.6kbp

32.2kbp

41398bp

30519bp

17244bp

12476bp

Nucleic Acids Res. 2007; 35:e16




Test with opgen technology

Data collection

About 1 Gb raw data from one device in 5 scans

(20 minutes)


Test with opgen technology

De novo Clustering


Test with opgen technology

Clusters in Opposite Orientation

150kb

150kb


Test with opgen technology

Histogram Plot

Consensus Map

Reference

Human genome data


Test with opgen technology

Cluster Mapped to human Chr 7

170kb



3048 cre linearization 90 efficiency
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


Test with opgen technology

Individual clone maps from clone mixture

Test with Ae.tauschii BAC clones


Test with opgen technology

Cone D1

HI064J24

BbvCI

BspQI

Start of

vector

Linearized by loxp

2kb extra

Asisi

site

BspQI

BbvCI

2kb extra

Linearized by AsiSi


Test with opgen technology

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