Difference between revisions of "Part:BBa K176003:Design"
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After eliminate the MCS, the lacZalpha part is identical to the N-terminal of the wild type gene(BBa_I732006 & BBa_I732005), and fused with the ccdB part (BBa_K145151) with a 3aa linker.
 
After eliminate the MCS, the lacZalpha part is identical to the N-terminal of the wild type gene(BBa_I732006 & BBa_I732005), and fused with the ccdB part (BBa_K145151) with a 3aa linker.
  
−
 
+
[[Image:K176003_MutiAlin.gif]]
  
 
===Source===
 
===Source===
  
−
PCR from the plasmid pluxCcdB3<cite>You</cite (a gift from Prof. Lingchong You>).
+
PCR from the plasmid pluxCcdB3<cite>You Balagadde</cite> (a gift from Prof. Lingchong You).
  
 
Forward Primer: GTTTCT TCTAG ATG accatgattacgg  attcactggccgtcgttttac
 
Forward Primer: GTTTCT TCTAG ATG accatgattacgg  attcactggccgtcgttttac
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Reverse Primer: GTTTCTTC CTGCAG CGGCCGC T ACTAGT A TTATTA tattccccagaacatcaggtta
 
Reverse Primer: GTTTCTTC CTGCAG CGGCCGC T ACTAGT A TTATTA tattccccagaacatcaggtta
  
−
Digest with XbaI and SpeI, ligate into pSB1A3.
+
Digest with XbaI and PstI, ligate into pSB1A3.
  
 
===References===
 
===References===
 
<biblio>
 
<biblio>
 
#You pmid=15064770
 
#You pmid=15064770
 +
#Balagadde pmid=18414488
 
</biblio>
 
</biblio>

Latest revision as of 12:34, 14 September 2009

lacZalpha-ccdB coding sequence


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
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 387


Design Notes

The part is similar to BBa_J32026 and the original coding sequence in pluxCcdB3, but the multiple cloning sites are eliminated in order to avoid site-specific mutagenesis. The multiple cloning sites are from lacZalpha in pZErO-2.

After eliminate the MCS, the lacZalpha part is identical to the N-terminal of the wild type gene(BBa_I732006 & BBa_I732005), and fused with the ccdB part (BBa_K145151) with a 3aa linker.

K176003 MutiAlin.gif

Source

PCR from the plasmid pluxCcdB3You Balagadde (a gift from Prof. Lingchong You).

Forward Primer: GTTTCT TCTAG ATG accatgattacgg attcactggccgtcgttttac

Reverse Primer: GTTTCTTC CTGCAG CGGCCGC T ACTAGT A TTATTA tattccccagaacatcaggtta

Digest with XbaI and PstI, ligate into pSB1A3.

References

<biblio>

  1. You pmid=15064770
  2. Balagadde pmid=18414488

</biblio>