How to do --- MSP or Clone-sequencing? - (Jul/11/2005 )
I intended to detect the methylation difference of CpG islands between case and control. And I have selected several candidate genes for study.
To get the methylation status of CpG islands in promoter regions before amplify sample for study, which method is preferable to use for selecting genes----MSP or Clone-sequencing?
As I know, MSP can only detect the on/off methyaltion status of CpG islands. How to differentiate between the complete methylation and part methylation by MSP?
In addition, if candidate genes in completely methylated or never methylated, are they be discard? Only partly methylated genes could be selected for further study?
Cloning and sequencing can provide information for methylation pattern in an allele and quantitative values. Does it appropriate for initial filter genes? How to exact its' best useness combined with MSP?
>>As I know, MSP can only detect the on/off methyaltion status of CpG islands. How to differentiate between the complete methylation and part methylation by MSP?
In MSP, you use two sets of primers, one is specific for methylated DNA (MSP) and the other for unmethylated DNA (USP). If your DNA is a mixture of methylated and unmethylated molecules, both sets of primer will show bands indicating partial methylation. Here, Partial means not all cells are equally methylated. If you talking about partial methylation along a CpG island (or regional methylation) of a single DNA molecule, you have to use multiple primer sets to map the status of different CpG sites.
>>In addition, if candidate genes in completely methylated or never methylated, are they be discard? Only partly methylated genes could be selected for further study?
Your goal is to find the difference between case and control.
>>Cloning and sequencing can provide information for methylation pattern in an allele and quantitative values. Does it appropriate for initial filter genes? How to exact its' best useness combined with MSP?
If you want to do something serious, go for BSP.