Part:BBa_J70012

A BioScaffold Part (Uses PsrI), Protein Head Remover see Part Design page

When placed in a BioBrick assembly and cut with the restriction enzyme PsrI, this part will direct the restriction enzyme to cut outside the BioBrick scar

still in testing, uses PsrI (2) AscI (2) MabI (2), currently in pSB1AK3 vector-> now being moved to pSB2K4 (it's final destination)

See part design page or BBFRFC15, for details about use

Notes:

We now have a sequenced version of the part "J70012final-pSB2K4" (from J70012 1b in DNA sequence page) and are testing the functionality of the part (whether it can be used in a "BioScaffold parts" context)

This part can be made with the following oligos, which can be annealed and placed into an EcoRI/SpeI cut vector:

Version 1.1 Oligos: These oligos produced a large number of transformants on the first try and all were correct.
For a future version, one may want to move the MabI sites closer to the center like in J70010.  

 EcoRI NotI     XbaI                                                           SpeI
  AATTCGCGGCCGCTTCTAGAG AACACCTGGTACAGCAGG |  TGGCGCGCCGGCGCGCCAG CTGGTGAACACCAGGTA (ctag)
      G                                                          |               at  gatc
 gaattcgcggccgcttctagag                                                          ta  ctagtagcggccgctgcag
 BioBricks Prefix                                                                BioBricks Suffix
Forward 1.1 (TM1=77.5): AATTCGCGGCCGCTTCTAGAGAACACCTGGTACAGCAGG
Forward 2.1 (TM1=66.2, TM2=47.1): TGGCGCGCCGGCGCGCCAGCTGGTGAACACCAGGTA
Reverse 1.1 (TM1=66.2, TM2=77.5): CTGGCGCGCCGGCGCGCCACCTGCTGTACCAGGTGTTCTCTAGAAGCGGCCGCG
Reverse 2.1 (TM=47.1): CTAGTACCTGGTGTTCACCAG

This part is currently being placed into the vector pSB2K4, which will allow hierarchical manipulations of the part.

3/18/9
We now have a sequenced version of the part "J70012final-pSB2K4" and are testing the functionality of the 
part (whether it can be used in a "BioScaffold parts" context) 


  AATTCGCG GCCGC^TTCTAGAG AACACCTGGTACAGCAGG |  TGGCGCGCCGGCGCGCCAG CTGGTGAACACCAGGtactagtagc ggccg^ctgcag
  TTAAGCGC^CGGCG AAGATCTC                                                          atgatcatcg^ccggc gacgtc

Potential test insert part 1:
(GCCGC)TGGCGCGCCGGCGCGCCAG GGCCG
 CGGCG ACCGCGCGGCCGCGCGGTC 
Forward TPJ70012.1.1.F (TM=66.2): TGGCGCGCCGGCGCGCCAGGGCCG
Reverse TPJ70012.1.1.R (TM=66.2): CTGGCGCGCCGGCGCGCCAGCGGC

Potential test insert part 0 (the null part):
(GCCGC)TTCTAGAGtactagtagc ggccg
 CGGCG AAGATCTCatgatcatcg
Forward TPJ70012.0.1.F (TM=43.5): TTCTAGAGtactagtagcggccg
Reverse TPJ70012.0.1.R (TM=43.5): gctactagtaCTCTAGAAGCGGC

3/31/9

AATTCGCG GCCGC^TTCTAGAG AACACCTGGTACAGCAGG |  TGGCGCGCCGGCGCGCCAG CTGGTGAACACCAGGtactagtagc ggccg^ctgcag
Make PCR part 
                     ag aacacctggtacagcagg    tgatgctagctagat
                                      62.2
                                                 ATGAACGATGCAGATG CTGGTGAACACCAGGtactag
                                                                 62.9
                                                                       
J70012.r.pcr reverse oligo: atctagctagcatcacctgctgtaccaggtgttct (21 bp beyond PsrI recognition site)
J70012.f.pcr forward oligo: ATGAACGATGCAGATG CTGGTGAACACCAGGtactag (21 bp beyond PsrI recognition site)

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 25
  
• 1000
  COMPATIBLE WITH RFC[1000]