LOCUS       sequence-371333-        5640 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 5640)
  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 5640)
  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-HALO-Hygromycin
FEATURES             Location/Qualifiers
     source          1..5640
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(31..459)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(141..162)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     353..372
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     586..608
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     600..617
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     601..617
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     627..646
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        627..645
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     670..686
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     671..687
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             736..825
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     CDS             856..1746
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     gene            1804..3379
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     promoter        1804..2147
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             2148..3176
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             2148..3176
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     terminator      3182..3379
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(3453..3473)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3453..3471)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3492..3508)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3492..3508)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3505..3527)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3516..3532
                     /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(3540..3546,3547..3564,3565..3570))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3585..3606
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3723..3740)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3894..4482)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3974..3993)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4653..5444,5445..5513))
                     /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     5276..5295
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5514..5618)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cctaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct
       61 cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg
      121 agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact
      181 ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac
      241 cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga
      301 gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga
      361 aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca
      421 ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc ccattcgcca ttcaggctgc
      481 gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag
      541 ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt
      601 gtaaaacgac ggccagtgag cgcgcgtaat acgactcact atagggcgaa ttgggtaccg
      661 ggccccccct cgaggtcgac ggtatcgata agcttgatat cgggtcgacg gatccccggg
      721 ttaattaaca tcttttaccc atacgatgtt cctgactatg cgggctatcc ctatgacgtc
      781 ccggactatg caggatccta tccatatgac gttccagatt acgctgctca gtgcggttca
      841 ggtggatctg gttctatggc agaaatcggt actggctttc cattcgaccc gcattatgtg
      901 gaagtcctgg gcgagagaat gcattacgtt gacgtgggtc cgagagatgg aactccggtc
      961 ctttttctgc acgggaatcc tacaagctct tatgtttggc gcaatatcat ccctcatgta
     1021 gctccgacgc atcgctgtat tgcgccggac ctgattggta tgggaaaatc tgataaacca
     1081 gacctgggtt actttttcga tgatcatgtg cgtttcatgg atgccttcat tgaggcatta
     1141 gggcttgaag aagtcgtctt ggtgattcat gattggggct cagctctggg atttcactgg
     1201 gctaaaagaa atcctgaacg cgtaaaaggc atcgcgttta tggagttcat tcgtccaatt
     1261 ccgacttggg atgaatggcc tgagttcgcg agagaaacat ttcaagcatt tcgcacgacc
     1321 gatgtaggcc ggaagttaat catcgatcag aatgtcttta tcgaagggac attgccgatg
     1381 ggagtcgttc gtccgttaac agaagtcgaa atggatcact atagagaacc ttttcttaat
     1441 cctgtggaca gagagccgct gtggcggttt ccgaacgaac tgccgattgc aggcgagcct
     1501 gctaacattg tagcgctggt tgaagagtat atggattggc ttcatcagtc tccagttccg
     1561 aagttattgt tttggggtac gcctggcgtg cttattccac cggccgaagc ggcacgtttg
     1621 gcaaaaagcc tgccaaattg caaagccgtt gacattggcc ctggacttaa cttgcttcaa
     1681 gaggataacc cggacttaat cgggagcgaa attgcccggt ggctttctac cttagaaatc
     1741 agcggctaat atataactat ctagaaataa agagtatcat ctttcaaagg tcacccggcc
     1801 agcgacatgg aggcccagaa taccctcctt gacagtcttg acgtgcgcag ctcaggggca
     1861 tgatgtgact gtcgcccgta catttagccc atacatcccc atgtataatc atttgcatcc
     1921 atacattttg atggccgcac ggcgcgaagc aaaaattacg gctcctcgct gcggacctgc
     1981 gagcagggaa acgctcccct cacagacgcg ttgaattgtc cccacgccgc gcccctgtag
     2041 agaaatataa aaggttagga tttgccactg aggttcttct ttcatatact tccttttaaa
     2101 atcttgctag gatacagttc tcacatcaca tccgaacata aacaaccatg ggtaaaaagc
     2161 ctgaactcac cgcgacgtct gtcgagaagt ttctgatcga aaagttcgac agcgtctccg
     2221 acctgatgca gctctcggag ggcgaagaat ctcgtgcttt cagcttcgat gtaggagggc
     2281 gtggatatgt cctgcgggta aatagctgcg ccgatggttt ctacaaagat cgttatgttt
     2341 atcggcactt tgcatcggcc gcgctcccga ttccggaagt gcttgacatt ggggaattca
     2401 gcgagagcct gacctattgc atctcccgcc gtgcacaggg tgtcacgttg caagacctgc
     2461 ctgaaaccga actgcccgct gttctgcagc cggtcgcgga ggccatggat gcgatcgctg
     2521 cggccgatct tagccagacg agcgggttcg gcccattcgg accgcaagga atcggtcaat
     2581 acactacatg gcgtgatttc atatgcgcga ttgctgatcc ccatgtgtat cactggcaaa
     2641 ctgtgatgga cgacaccgtc agtgcgtccg tcgcgcaggc tctcgatgag ctgatgcttt
     2701 gggccgagga ctgccccgaa gtccggcacc tcgtgcacgc ggatttcggc tccaacaatg
     2761 tcctgacgga caatggccgc ataacagcgg tcattgactg gagcgaggcg atgttcgggg
     2821 attcccaata cgaggtcgcc aacatcttct tctggaggcc gtggttggct tgtatggagc
     2881 agcagacgcg ctacttcgag cggaggcatc cggagcttgc aggatcgccg cggctccggg
     2941 cgtatatgct ccgcattggt cttgaccaac tctatcagag cttggttgac ggcaatttcg
     3001 atgatgcagc ttgggcgcag ggtcgatgcg acgcaatcgt ccgatccgga gccgggactg
     3061 tcgggcgtac acaaatcgcc cgcagaagcg cggccgtctg gaccgatggc tgtgtagaag
     3121 tactcgccga tagtggaaac cgacgcccca gcactcgtcc gagggcaaag gaataatcag
     3181 tactgacaat aaaaagattc ttgttttcaa gaacttgtca tttgtatagt ttttttatat
     3241 tgtagttgtt ctattttaat caaatgttag cgtgatttat attttttttc gcctcgacat
     3301 catctgccca gatgcgaagt taagtgcgca gaaagtaata tcatgcgtca atcgtatgtg
     3361 aatgctggtc gctatactgc tgtcgattcg atactaacgc cgccactagt tctagagcgg
     3421 ccgccaccgc ggtggagctc cagcttttgt tccctttagt gagggttaat tgcgcgcttg
     3481 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     3541 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     3601 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     3661 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     3721 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     3781 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     3841 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     3901 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     3961 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     4021 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     4081 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     4141 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     4201 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     4261 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     4321 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     4381 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     4441 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     4501 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     4561 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     4621 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     4681 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     4741 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     4801 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     4861 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     4921 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     4981 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     5041 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     5101 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     5161 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     5221 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     5281 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     5341 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     5401 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     5461 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     5521 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     5581 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
//
