Methods And Compositions For High-Throughput Bisulphite Dna-Sequencing And Utilities
The Invention Relates To Novel Methods And Compositions To Produce Dna Templates Suitable For Chemical Modifications And High-Throughput Dna-Sequencing. A Method Of The Invention Relates To A Dna Adaptor Design Where Constituent Deoxycytosines Are Substituted With 5-Methyl-Deoxycytosines Rendering The Resulting Adaptor Resistant To Bisulphite Mediated Deamination. When Said Adaptor Is Ligated Onto Double Stranded Dna Template, Subsequent Dna Denaturation And Bisulphite Treatment Deaminates Template Dna Deoxycytosine Differentially To Deoxyuraeil Whilst The 5-Methyl-Deoxycytosines Of The Ligated Adaptor Resist Chemical Conversion Resulting In The Adaptor Sequence Remaining Unaltered. Both Strands Of Bisulphite Treated Dna Can Thus Be Amplified With A Single Primer Set That Hybridizes To The Unaltered Adaptor Sequence. The Invention Also Relates To Methods To Produce Control Template Of A Defined Methylation Composition To Optimize Conditions For The Bisulphite Reaction. In A Preferred Embodiment, The Present Invention Can Be Used To Produce Templates Suitable For Genome-Wide Bisulphite-Dna Sequencing Using Conventional, Solexa(Tm), Solid(Tm) Or 454(Tm)-Type Dna Sequencing Platforms To Study Dna Methylation.
Patent Number: US20090047680A1
Application Number: US2008192393A
Inventor: LOK, SI
Priority Date: 15 Aug 2007
Priority Number: US20090047680A1
Application Date: 15 Aug 2008
Publication Date: 19 Feb 2009
IPC Current: C12Q000168 | C12P001934
US Class: 43500614 | 435006 | 4350912
Title: METHODS AND COMPOSITIONS FOR HIGH-THROUGHPUT BISULPHITE DNA-SEQUENCING AND UTILITIES
Usefulness: METHODS AND COMPOSITIONS FOR HIGH-THROUGHPUT BISULPHITE DNA-SEQUENCING AND UTILITIES
Summary: The method is useful for determining the epigenomic status of various DNA target populations, specifically for determining the methylation of deoxycytosine at the 5-carbon position. The methods are also used for producing hemi-methylated DNA control templates and methylated DNA control templates which are used to monitor bisulfite reaction efficiency (all claimed). The templates are used for genome-wide bisulfite-DNA sequencing using conventional platforms to study DNA methylation.
Novelty: Determining the epigenomic status of target DNA population comprises fragmenting the DNA population, ligating modified adaptors to target DNA, subjecting to chemical treatment, amplifying the mixture, and sequencing the amplified DNA
生物醫學
DNA /基因工程
香港
