LOCUS       sequence-371326-        4770 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Integration cassette to create C-terminal fusions for fluorescent 
            tagging.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4770)
  AUTHORS   Baek I, Le SN, Jeon J, Chun Y, Reed C, Buratowski S
  TITLE     A set of Saccharomyces cerevisiae integration vectors for 
            fluorescent dye labeling of proteins.
  JOURNAL   G3 (Bethesda). 2022 Aug 9. pii: 6659097. doi: 
            10.1093/g3journal/jkac201.
  PUBMED    35944214
REFERENCE   2  (bases 1 to 4770)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "G3 
            (Bethesda). 2022 Aug 9. pii: 6659097. doi: 
            10.1093/g3journal/jkac201."
COMMENT     Sequence Label: pBS-SKII-3XHA-eDHFR-NAT
FEATURES             Location/Qualifiers
     source          1..4770
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(19..474)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(156..177)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     368..387
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     601..623
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     615..632
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     616..632
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     642..661
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        642..660
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     685..701
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     686..702
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             751..840
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     CDS             871..1350
                     /codon_start=1
                     /gene="folA"
                     /product="E. coli dihydrofolate reductase"
                     /label=ecDHFR
                     /translation="MISLIAALAVDRVIGMENAMPWNLPADLAWFKRNTLNKPVIMGRH
                     TWESIGRPLPGRKNIILSSQPGTDDRVTWVKSVDEAIAACGDVPEIMVIGGGRVYEQFL
                     PKAQKLYLTHIDAEVEGDTHFPDYEPDDWESVFSEFHDADAQNSHSYCFEILERR"
     gene            1405..2524
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     promoter        1405..1748
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1749..2321
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     terminator      2327..2524
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(2598..2618)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2598..2616)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2637..2653)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2637..2653)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2650..2672)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2661..2677
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(2685..2691,2692..2709,2710..2715))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2730..2751
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2868..2885)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3039..3627)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3119..3138)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3798..4589,4590..4658))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4421..4440
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4659..4763)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ccccgaaaag tgccacctaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt
       61 tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat
      121 caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat
      181 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac
      241 tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc
      301 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga
      361 gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca
      421 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcccatt
      481 cgccattcag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct tcgctattac
      541 gccagctggc gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt
      601 cccagtcacg acgttgtaaa acgacggcca gtgagcgcgc gtaatacgac tcactatagg
      661 gcgaattggg taccgggccc cccctcgagg tcgacggtat cgataagctt gatatcgggt
      721 cgacggatcc ccgggttaat taacatcttt tacccatacg atgttcctga ctatgcgggc
      781 tatccctatg acgtcccgga ctatgcagga tcctatccat atgacgttcc agattacgct
      841 gctcagtgcg gttcaggtgg atctggttct atgatcagtc tgattgcggc gttagcggta
      901 gatcgcgtta tcggcatgga aaacgccatg ccgtggaacc tgcctgccga tctcgcctgg
      961 tttaaacgca acaccttaaa taaacccgtg attatgggcc gccatacctg ggaatcaatc
     1021 ggtcgtccgt tgccaggacg caaaaatatt atcctcagca gtcaaccggg tacggacgat
     1081 cgcgtaacgt gggtgaagtc ggtggatgaa gccatcgcgg cgtgtggtga cgtaccagaa
     1141 atcatggtga ttggcggcgg tcgcgtttat gaacagttct tgccaaaagc gcaaaaactg
     1201 tatctgacgc atatcgacgc agaagtggaa ggcgacaccc atttcccgga ttacgagccg
     1261 gatgactggg aatcggtatt cagcgaattc cacgatgctg atgcgcagaa ctctcacagc
     1321 tattgctttg agattctgga gcggcggtaa tatataactg tctagaaata aagagtatca
     1381 tctttcaaag gtcacccggc cagcgacatg gaggcccaga ataccctcct tgacagtctt
     1441 gacgtgcgca gctcaggggc atgatgtgac tgtcgcccgt acatttagcc catacatccc
     1501 catgtataat catttgcatc catacatttt gatggccgca cggcgcgaag caaaaattac
     1561 ggctcctcgc tgcagacctg cgagcaggga aacgctcccc tcacagacgc gttgaattgt
     1621 ccccacgccg cgcccctgta gagaaatata aaaggttagg atttgccact gaggttcttc
     1681 tttcatatac ttccttttaa aatcttgcta ggatacagtt ctcacatcac atccgaacat
     1741 aaacaaccat gggtaccact cttgacgaca cggcttaccg gtaccgcacc agtgtcccgg
     1801 gggacgccga ggccatcgag gcactggatg ggtccttcac caccgacacc gtcttccgcg
     1861 tcaccgccac cggggacggc ttcaccctgc gggaggtgcc ggtggacccg cccctgacca
     1921 aggtgttccc cgacgacgaa tcggacgacg aatcggacga cggggaggac ggcgacccgg
     1981 actcccggac gttcgtcgcg tacggggacg acggcgacct ggcgggcttc gtggtcgtct
     2041 cgtactccgg ctggaaccgc cggctgaccg tcgaggacat cgaggtcgcc ccggagcacc
     2101 gggggcacgg ggtcgggcgc gcgttgatgg ggctcgcgac ggagttcgcc cgcgagcggg
     2161 gcgccgggca cctctggctg gaggtcacca acgtcaacgc accggcgatc cacgcgtacc
     2221 ggcggatggg gttcaccctc tgcggcctgg acaccgccct gtacgacggc accgcctcgg
     2281 acggcgagca ggcgctctac atgagcatgc cctgccccta atcagtactg acaataaaaa
     2341 gattcttgtt ttcaagaact tgtcatttgt atagtttttt tatattgtag ttgttctatt
     2401 ttaatcaaat gttagcgtga tttatatttt ttttcgcctc gacatcatct gcccagatgc
     2461 gaagttaagt gcgcagaaag taatatcatg cgtcaatcgt atgtgaatgc tggtcgctat
     2521 actgctgtcg attcgatact aacgccgcca ctagttctag agcggccgcc accgcggtgg
     2581 agctccagct tttgttccct ttagtgaggg ttaattgcgc gcttggcgta atcatggtca
     2641 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     2701 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     2761 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
     2821 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     2881 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     2941 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     3001 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     3061 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     3121 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     3181 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     3241 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     3301 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     3361 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     3421 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     3481 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     3541 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     3601 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     3661 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     3721 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     3781 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     3841 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     3901 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     3961 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     4021 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     4081 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     4141 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     4201 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     4261 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     4321 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     4381 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     4441 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     4501 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     4561 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     4621 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     4681 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     4741 aataaacaaa taggggttcc gcgcacattt
//
