Difference between revisions of "Part:BBa J31002:Design"

 
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===Design Notes===
 
===Design Notes===
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Kan was PCR-amplified (from pSB1AK3) with primers designed to introduce a SpeI site to the left and an XbaI site to the right, reversing the normal order of these two BioBrick cut sites. The PCR amplicon was digested with SpeI and XbaI and ligated into S/X cut pSB1A2. After cloning, we used a S/X digest to check for insertion in the desired orientation (reverse). Clones where Kan inserted in the wrong way failed to excise (due to S/X mixed sites) and clones where Kan inserted in the desired orientation excised successfully (restoration of S and X sites).
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===Source===
 
===Source===

Latest revision as of 00:56, 29 October 2006


kanamycin resistance backwards (KanB) [cf. BBa_J23012 & BBa_J31003]


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

Kan was PCR-amplified (from pSB1AK3) with primers designed to introduce a SpeI site to the left and an XbaI site to the right, reversing the normal order of these two BioBrick cut sites. The PCR amplicon was digested with SpeI and XbaI and ligated into S/X cut pSB1A2. After cloning, we used a S/X digest to check for insertion in the desired orientation (reverse). Clones where Kan inserted in the wrong way failed to excise (due to S/X mixed sites) and clones where Kan inserted in the desired orientation excised successfully (restoration of S and X sites).

Source

pSB1AK3

References