Difference between revisions of "Part:BBa K174000:Design"

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==Construction==
 
==Construction==
Treat Plasmid part BBa_J04450 (pSB1AT3) and our construct with EcoRI and SpeI
+
Plasmid part BBa_J04450 (pSB1AT3) and our construct were treated with EcoRI and SpeI
  
 
[[Image:Newcastle_2009_sspB_1.png‎]]
 
[[Image:Newcastle_2009_sspB_1.png‎]]
  
Ligation of the backbone and the sspB insert
+
The backbone and the sspB insert were then ligated
  
 
[[Image:Newcastle_2009_sspB_2.png‎]]
 
[[Image:Newcastle_2009_sspB_2.png‎]]

Revision as of 19:33, 26 September 2009

498 bp long sspB CDS is amplified by PCR with dangling end primers with EcoRI-NotI-XbaI restriction site at 5’ and SpeI at 3’ and inserted into a Biobrick compatible vector. The sequence is taken from E. coli strain DH5 alpha. Genbank accession number for E. coli MG1655 strain is NC_000913.2

Forward primer used: GATCTG-GAATTCGCGGCCGCTTCTAG-ATGGATTTGTCACAGCTAAC (Clamp sequence - Standard Biobrick prefix - first 20 base from the Biobrick)

Reverse primer used: TGTGAC-ACTAGTA-TTACTTCACAACGCGTAATGC (Clamp sequence - SpeI site - last 21 base from the Biobrick)

Construction

Plasmid part BBa_J04450 (pSB1AT3) and our construct were treated with EcoRI and SpeI

Newcastle 2009 sspB 1.png

The backbone and the sspB insert were then ligated

Newcastle 2009 sspB 2.png