Hello everyone.
I'm currently trying to figure out how solid-phase/bridge-amplification works. Something still bothering me is, that the amplification step will create both the forward and the complementary strand.
According to slides 19-22 of http://sfgf.stanford.edu/.../general_technology.pdf the reverse strands will be removed (for single read).
But how does one know which one is the forward strand or the reversed strand?
Because we can't rely on the adapters, for identification!
As for slide 15 http://www.broadinstitute.org/.../SamplePrepSlides.pdf
the adapters will be added to the ends of the fragment, but we don't know which ones.
If for example CCCC is the sequence (of the forward strand), A1/A2 are the adapters and A1'/A2' their complements, our template might look like
5': A1 - T - CCCC - A - A2 : 3'
3': A1'- A - GGGG - T - A2' : 5'
or
5': A2'- T - CCCC - A - A1' :3'
3': A2 - A - GGGG - T - A1 :5'
Here A1 marks the forward strand in the first case, but the reversed strand in the second case.
Perhaps you can help me with that question.
I'm currently trying to figure out how solid-phase/bridge-amplification works. Something still bothering me is, that the amplification step will create both the forward and the complementary strand.
According to slides 19-22 of http://sfgf.stanford.edu/.../general_technology.pdf the reverse strands will be removed (for single read).
But how does one know which one is the forward strand or the reversed strand?
Because we can't rely on the adapters, for identification!
As for slide 15 http://www.broadinstitute.org/.../SamplePrepSlides.pdf
the adapters will be added to the ends of the fragment, but we don't know which ones.
If for example CCCC is the sequence (of the forward strand), A1/A2 are the adapters and A1'/A2' their complements, our template might look like
5': A1 - T - CCCC - A - A2 : 3'
3': A1'- A - GGGG - T - A2' : 5'
or
5': A2'- T - CCCC - A - A1' :3'
3': A2 - A - GGGG - T - A1 :5'
Here A1 marks the forward strand in the first case, but the reversed strand in the second case.
Perhaps you can help me with that question.
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