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Restriction enzyme based whole genome sequencing

A restriction enzyme and restriction technology, applied in the field of sequencing of biological genomes or parts thereof, can solve the problems of fierce competition, heavy, expensive draft genome sequences, etc.

Inactive Publication Date: 2012-09-05
KEYGENE NV
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  • Summary
  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

[0008] Despite all the developments in high-throughput sequencing, determining draft genome sequences with high accuracy is still considered expensive and onerous, and there is intense competition in the market

Method used

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  • Restriction enzyme based whole genome sequencing
  • Restriction enzyme based whole genome sequencing
  • Restriction enzyme based whole genome sequencing

Examples

Experimental program
Comparison scheme
Effect test

Embodiment

[0195] WGPS embodiment

[0196] The melon BAC library superset 24 is presented together with WGP data from superset 24 to demonstrate successful WGPS on the BAC set by splicing GA paired-end reads linked to enzyme sites.

[0197] 1. Wetlab method

[0198] The method contains the following steps:

[0199] - Digest (individual) BAC pool DNAs using a single enzyme (EcoRI).

[0200] - Ligation of pool-specific EcoRI compatible adapters containing P5 amplification, sequence primer 1 and pool-specific identifier sequences.

[0201] - (optionally pooling RL products from supersets that will be sequenced in a single pass on e.g. an Illumina Genome Analyzer. This is up to a maximum of different set-specific identifiers used in the previous ligation step)

[0202] - Fragmentation of the adapter ligated products into products with a size in the range of 100-1000 bp.

[0203] Fragmented adapter-ligated restriction fragments were blunt-ended and a single A-nt was added to the fragmente...

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Abstract

A method for de novo whole genome sequencing based on a (sequence-based) physical map of a DNA sample clone bank based on end-sequencing tagged adapter-ligated restriction fragments, in combination with sequencing adapter-ligated restriction fragments of the DNA sample wherein the recognition sequence of the restriction enzyme used in the generation of the physical map is identical to at least part of the recognition sequence of the restriction enzyme used in the generation of the DNA sample.

Description

technical field [0001] The present invention relates to methods and strategies for the efficient generation of whole genome sequences or parts thereof using high throughput sequencing. The present invention relates to large-scale nucleic acid sequencing and in particular to methods for the sequencing of biological genomes or parts thereof. The present invention relates to an improved strategy for determining the sequence of preferably complex (ie large) genomes based on the use of high-throughput sequencing technologies. Background technique [0002] The goal of many sequencing projects is to determine for the first time the entire genome sequence of a target organism (de novo draft genome sequencing). Having a draft genome sequence at hand enables the identification of biologically useful genetic information, for example for identifying the origin of genetic variation between species or between individuals of the same species. Therefore, techniques that allow de novo dete...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
CPCC12Q1/6869C12Q2525/191C12Q2521/301
Inventor R·C·J·赫格斯M·J·T·凡艾克
Owner KEYGENE NV
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