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I was wondering if someone could please comment on how the SOLiD-SAGE tags are produced in relation to a reference sequence from RefSeq? I have seen their protocol at http://tools.invitrogen.com/content/...D_SAGE_man.pdf and page 2 explains how the tags are generated, however it is not clear to me which strand and orientation the final sequenced tags will be when mapped on to a reference sequence.
I have used SOLiD-SAGE analysis software v1.10 to map the tags on a reference sequence database and found that the tags are reverse complements of the actual reference tags, not only in our dataset but also in the test data provided with the SOLiD-SAGE software. When we contacted a local lifetech support person we were told that the sequences are obtained in 3’ to 5’ direction but tehse should not be the reverse complements of actual reference sequence. This has really confused us.
I was wondering if any of you have tried to map the SOLiD-SAGE tags using any other tool except SOLiD-SAGE software v1.10 (e.g. BWA or PerM). One difference between SOLiD-SAGE software and PerM, for example, is that SOLiD-SAGE software takes in a reference fasta file for mapping and makes its own virtual tags, while we make virtual tags ourselves before using PerM to map SOLiD-tags on the virtual tags. Hence it is important that we get the right direction and orientation of the virtual tags for mapping SOLiD tags. I would appreciate if someone could comment on this pelase.
Cheers
I was wondering if someone could please comment on how the SOLiD-SAGE tags are produced in relation to a reference sequence from RefSeq? I have seen their protocol at http://tools.invitrogen.com/content/...D_SAGE_man.pdf and page 2 explains how the tags are generated, however it is not clear to me which strand and orientation the final sequenced tags will be when mapped on to a reference sequence.
I have used SOLiD-SAGE analysis software v1.10 to map the tags on a reference sequence database and found that the tags are reverse complements of the actual reference tags, not only in our dataset but also in the test data provided with the SOLiD-SAGE software. When we contacted a local lifetech support person we were told that the sequences are obtained in 3’ to 5’ direction but tehse should not be the reverse complements of actual reference sequence. This has really confused us.
I was wondering if any of you have tried to map the SOLiD-SAGE tags using any other tool except SOLiD-SAGE software v1.10 (e.g. BWA or PerM). One difference between SOLiD-SAGE software and PerM, for example, is that SOLiD-SAGE software takes in a reference fasta file for mapping and makes its own virtual tags, while we make virtual tags ourselves before using PerM to map SOLiD-tags on the virtual tags. Hence it is important that we get the right direction and orientation of the virtual tags for mapping SOLiD tags. I would appreciate if someone could comment on this pelase.
Cheers