Difference between revisions of "Part:BBa K2847002:Design"

 
 
Line 7: Line 7:
  
 
===Design Notes===
 
===Design Notes===
−
PCR amplification with primers: F-CAGGTTTGTGCCAATACCAG and R-CAGGTGGCACTTTTCGGG
+
This part was PCR amplified with primers:
 +
 
 +
Primer_F: TATCTGCAGCAGGTTTGTGCCAATACCAG
 +
 
 +
Primer_R: TATGAATTCCAGGTGGCACTTTTCGGG
 +
 
 +
 
 +
using pE-FLP (Addgene Plasmid #45978) as a matrix. In order to obtain the T-sensitive SC101 origin of replication one can use the following primer pair:
 +
 
 +
Primer_F: CAGGTTTGTGCCAATACCAG
 +
 
 +
Primer_R CAGCGATTTGCCCGATTGC
 +
 
 +
 
 +
The required 24-bp overhangs for Aqua cloning based integration into pSB1 vector backbones should have the following sequences:
 +
 
 +
cggccgcaatcgggcaaatcgctg
 +
 
 +
tctactggtattggcacaaacctg
 +
 
 +
 
 +
 
  
  

Latest revision as of 19:01, 17 October 2018


pSC101 ori + RepA (thermosensitive mutant) + AmpR gene


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
    Illegal SpeI site found at 1163
  • 12
    INCOMPATIBLE WITH RFC[12]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
    Illegal SpeI site found at 1163
    Illegal NotI site found at 1482
  • 21
    INCOMPATIBLE WITH RFC[21]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
  • 23
    INCOMPATIBLE WITH RFC[23]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
    Illegal SpeI site found at 1163
  • 25
    INCOMPATIBLE WITH RFC[25]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
    Illegal SpeI site found at 1163
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Plasmid lacks a prefix.
    Plasmid lacks a suffix.
    Illegal BsaI.rc site found at 2128


Design Notes

This part was PCR amplified with primers:

Primer_F: TATCTGCAGCAGGTTTGTGCCAATACCAG

Primer_R: TATGAATTCCAGGTGGCACTTTTCGGG


using pE-FLP (Addgene Plasmid #45978) as a matrix. In order to obtain the T-sensitive SC101 origin of replication one can use the following primer pair:

Primer_F: CAGGTTTGTGCCAATACCAG

Primer_R CAGCGATTTGCCCGATTGC


The required 24-bp overhangs for Aqua cloning based integration into pSB1 vector backbones should have the following sequences:

cggccgcaatcgggcaaatcgctg

tctactggtattggcacaaacctg




Source

Plasmid pE-FLP (Addgene Plasmid #45978)

References