BBa_K300001 construction intermediate (provided by Mr Gene)
Mr Gene synthesis service (www.mrgene.com).
This part has been provided by Mr Gene synthesis service and it has been used to construct BBa_K300001 plasmid.
Susanna Zucca, Federica Sampietro, Paolo Magni
This site can also be cut with PstI restriction enzyme.
Homologous region in S288C yeast genome (GAL1 ORF)
AvrII restriction enzyme site (XbaI, SpeI compatible overhang)
AvrII recognition site. Digestion with AvrII generates a compatible cohesive end to XbaI and SpeI.
MfeI restriction enzyme site (EcoRI compatible overhang)
MfeI recognition site. Digestion with MfeI yields a compatible cohesive end to EcoRI.
NheI restriction enzyme site (XbaI, SpeI compatible overhang)
NheI recognition site. Digestion with NheI generates a compatible cohesive end to XbaI and SpeI.
RBS.4 (weaker) -- derivative of BBa_0030
Weaker RBS based on Ron Weiss thesis. Strengths relative to <bb_part>BBa_B0030</bb_part>, <bb_part>BBa_B0031</bb_part>, <bb_part>BBa_B0032</bb_part>.
Varies from -6 to +1 region from original sequence to accomodate BioBricks suffix ("RBS-3" in figure 4-14 of thesis). No secondary structures are formed in the given RBS region. Users should check for secondary structures induced in the RBS by upstream and downstream elements in the +50 to -50 region, as such structures will greatly affect the strength of the RBS. Contact info for this part: Brian Chow
Vinay S Mahajan, Voichita D. Marinescu, Brian Chow, Alexander D Wissner-Gross and Peter Carr IAP, 2003.
[LoxP-KanMX-LoxP] excisable marker for yeast
This cassette is to be used in yeast. It is composed by the kanMX gene from the E. coli transposon Tn903 that confers the resistance to the aminoglycoside antibiotic G418 to yeast (commonly used at 200 ug/ml working concentration). This gene is expressed by the TEF2 promoter/RBS and terminated by the TEF2 transcriptional terminator from Ashbya gossypii filamentous fungus. The cassette is flanked by two loxP sites and for this reason it can be excised with Cre/loxP mediated marker removal recombination procedure. ADVANTAGES IN GENOME ENGINEERING APPLICATIONS: This part can be used to select positive integrants in the yeast genome and then it can be eliminated through Cre/loxP procedure. This enables the engineering of yeast genome in multiple steps, always using the same dominant selection marker.
The original sequence of loxP-KanMX-loxP from pUG6 vector has two NsiI restriction sites in kanR(Tn903) coding sequence and they have been removed in this part by changing codons. The TEF2 promoter has a XbaI and a PstI restriction sites, which have not been removed to avoid the change of the promoter strength. For this reason, this part is not fully compatible with RFC10 BioBrick Standard Assembly, but it can be assembled by using EcoRI-SpeI enzymes.
Lorenzo Pasotti, Giacomo Zambianchi, Federica Sampietro, Paolo Magni
Homologous region in S288C yeast genome (GAL10 ORF)
NsiI restriction enzyme site (PstI compatible overhang)
NsiI recognition site. Digestion with NsiI generates a compatible cohesive end to PstI.
Conversion of the iGEM parts registry to SBOL2.1