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 Ura3
| Types | DnaRegion |
| Roles | Plasmid_Backbone plasmid_vector |
| Sequences | BBa_K394002_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 Ura3 locus in S. cerevisiae. To transform S. cerevisiae the plasmid is cut with BbsI creating two non-sticky ends. The linear fragment then contains Ura3 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 Ura- 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 Ura3 homology VR rep (pMB1) (rev) ORI Tet-R AmpR (rev) VF2 Ura3 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 3387,3435 3435,3457 3381,3386 72,77 | engineered_region feature/BioBrick feature/conserved conserved_region primer_binding_site feature/primer_binding feature/cds CDS sequence_feature feature/misc CDS feature/cds feature/cds CDS feature/primer_binding primer_binding_site feature/conserved conserved_region feature/BioBrick engineered_region feature/misc sequence_feature feature/misc sequence_feature |