Koch Lab:Protocols/Sequences/PCR Primers
Our naming convention for plasmids is as follows:
plasmid name + forward or reverse primer + index of 5' match + label
For example, pRL574-R2008-bio is a primer that binds the reverse strand of pRL574, with it's 5' match at base 2008, and a biotin label (position of label not specified in name, but often it's at the 5' end).
Primers for Plasmid pRL574
Plasmid pRL574 is a plasmid out of the Landick lab that was developed for transcription studies. Because of their seminar work in single-molecule transcription studies, many other non-transcription-related single-molecule experiments evolved that use this plasmid. We have a bunch of PCR primers that work on this plasmid to create various sized segments.
pRL574-F853 or pRL574-F853-dig
This is the primer "P1" Steve used in grad school
5' GTAAAACGACGGCCAGTGAATTC 3' 23 nucleotides
pRL574-F834 or pRL574-F834-dig
Redesigned to substitute for F853. Designed to pair with pRL574-R2044; apparently works well with both R2044 and R2088
5' TTTTCCCAGTCACGACGTTG 3' 20 nucleotides Biopolymer calc predicted melting temperature: 59.4, mFold: 59.7 (50 mM NaCl, 10 uM oligo), no hairpins detected
pRL574-R2008 or pRL574-R2008-bio
This is the primer "P13" that Steve used in grad school
5' CACGTAAGGTTTCAGAGATATATGGG 3' 26 nucleotides
pRL574-R2044 or pRL574-R2044-bio
designed to pair with pRL574-F834, apparently works just as well with F-853 as R2008 does.
5' CTACCAGTGCGCTCAGACG 3' - or - 5'-bio-CTACCAGTGCGCTCAGACG 3' 19 nucleotides Biopolymer calc predicted melting temperature: 58.0, mFold: 62.2, no hairpins detected
Designed by Brandon beck for easier use during BstXI cutting (smaller fragment after cut)
5' GGGCCGTGATCCAGATCG 3'
pRL574-R5263 and pRL574-R5263-bio
5'- GGAAACAGCTATGACCATGATTAC - or - 5'-bio-GGAAACAGCTATGACCATGATTAC