Source:
Generated By: https://synbiohub.org/public/igem/igem2sbol/1
Created by: Brian Landry
Date created: 2010-10-19 11:00:00
Date modified: 2015-05-08 01:12:21
Plasmid for Chromosomal Integration in Yeast at His3
| Types | DnaRegion |
| Roles | plasmid_vector Plasmid_Backbone |
| Sequences | BBa_K394001_sequence (Version 1) |
Description
This part is designed to allow simple chromosomal integration of BioBrick plasmids. It is based off of the bacterial pSB1AT3 backbone allowing for normal DNA manipulation to occur in bacteria. However the BioBrick cut sites are flanked by homologous regions to the His3 locus in S. cerevisiae. To transform S. cerevisiae the plasmid is cut with BbsI creating two non-sticky ends. The linear fragment then contains His3 homologous sequences at its ends that can be used to chromosomally integrate the BioBrick plasmid into the yeast genome. Transformants can then be selected for using His- medium.Notes
NoneSource
This part was derived from plasmid pSB1AT3. Two successive rounds of PCR inserted a BbsI cut site and 40 bp homologous region in before the prefix and after the suffix.The following Primers were used:
Forward 1 GCACAGAACAATAACCTGCTGGAAACGAAGATAAATCgaagacGATTACTTCGCGTTATGCAGGC
Reverse 1 GCATCTTCTCAAATATGCTTCCCAGCCTGCTTATCcttctgAAATTCTGCCTCGTGATACGCC
Forward 2 tactagtagcggccgctgcagTCTTAACCCAACTGCACAGAACAATAACCTGCTGGAAACG
Reverse 2 ctctagaagcggccgcgaattcTTAGTATTGCTGGCCGCATCTTCTCAAATATGCTTCCCAGCC
| Sequence Annotation | Location | Component / Role(s) |
| BioBrick Suffix His3 homology VR rep (pMB1) (rev) ORI Tet-R AmpR (rev) VF2 His3 homology BioBrick Prefix BbsI Cut Site BbsI Cut Site | 1,21 22,69 157,176 261,875 276,276 1127,2317 2324,3184 3307,3326 3386,3435 3435,3457 71,76 3379,3385 | feature/BioBrick engineered_region feature/conserved conserved_region feature/primer_binding primer_binding_site CDS feature/cds feature/misc sequence_feature feature/cds CDS CDS feature/cds primer_binding_site feature/primer_binding conserved_region feature/conserved engineered_region feature/BioBrick sequence_feature feature/misc feature/misc sequence_feature |