LOCUS       pBS-SKII-3XHA-fS        5295 bp DNA     circular SYN 27-JUN-2019
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5295)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 14, 2022 from SnapGene 6.1.0
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5295
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(3..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     670..686
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             735..824
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     CDS             855..1400
                     /codon_start=1
                     /product="human O6-alkylguanine-DNA-alkyltransferase 
                     variant that forms covalent bonds with benzylguanine 
                     derivatives"
                     /label=SNAP-tag(R)
                     /note="fast-reacting variant, optimized for mammalian 
                     expression"
                     /translation="MDKDCEMKRTTLDSPLGKLELSGCEQGLHRIIFLGKGTSAADAVE
                     VPAPAAVLGGPEPLMQATAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLK
                     VVKFGEVISYSHLAALAGNPAATAAVKTALSGNPVPILIPCHRVVQGDLDVGGYEGGLA
                     VKEWLLAHEGHRLGKPGLG"
     gene            1458..3033
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     promoter        1458..1801
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1802..2830
                     /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      2836..3033
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     promoter        complement(3107..3125)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3146..3162)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3170..3186
                     /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(3194..3224)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3239..3260
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(3548..4136)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4307..5167)
                     /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"
     promoter        complement(5168..5272)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tgggtaccgg
      661 gccccccctc gaggtcgacg gtatcgataa gcttgatatc gggtcgacgg atccccgggt
      721 taattaacat cttttaccca tacgatgttc ctgactatgc gggctatccg tatgacgtcc
      781 cggactatgc aggatcctat ccatatgacg ttccagatta cgctgctcag tgcggttcag
      841 gtggatctgg ttctatggac aaagactgcg aaatgaagcg caccaccctg gatagccctc
      901 tgggcaagct ggaactgtct gggtgcgaac agggcctgca ccgtatcatc ttcctgggca
      961 aaggaacatc tgccgccgac gccgtggaag tgcctgcccc agccgccgtg ctgggcggac
     1021 cagagccact gatgcaggcc accgcctggc tcaacgccta ctttcaccag cctgaggcca
     1081 tcgaggagtt ccctgtgcca gccctgcacc acccagtgtt ccagcaggag agctttaccc
     1141 gccaggtgct gtggaaactg ctgaaagtgg tgaagttcgg agaggtcatc agctacagcc
     1201 acctggccgc cctggccggc aatcccgccg ccaccgccgc cgtgaaaacc gccctgagcg
     1261 gaaatcccgt gcccattctg atcccctgcc accgggtggt gcagggcgac ctggacgtgg
     1321 ggggctacga gggcgggctc gccgtgaaag agtggctgct ggcccacgag ggccacagac
     1381 tgggcaagcc tgggctgggt taatatataa ctgtctagaa ataaagagta tcatctttca
     1441 aaggtcaccc ggccagcgac atggaggccc agaataccct ccttgacagt cttgacgtgc
     1501 gcagctcagg ggcatgatgt gactgtcgcc cgtacattta gcccatacat ccccatgtat
     1561 aatcatttgc atccatacat tttgatggcc gcacggcgcg aagcaaaaat tacggctcct
     1621 cgctgcggac ctgcgagcag ggaaacgctc ccctcacaga cgcgttgaat tgtccccacg
     1681 ccgcgcccct gtagagaaat ataaaaggtt aggatttgcc actgaggttc ttctttcata
     1741 tacttccttt taaaatcttg ctaggataca gttctcacat cacatccgaa cataaacaac
     1801 catgggtaaa aagcctgaac tcaccgcgac gtctgtcgag aagtttctga tcgaaaagtt
     1861 cgacagcgtc tccgacctga tgcagctctc ggagggcgaa gaatctcgtg ctttcagctt
     1921 cgatgtagga gggcgtggat atgtcctgcg ggtaaatagc tgcgccgatg gtttctacaa
     1981 agatcgttat gtttatcggc actttgcatc ggccgcgctc ccgattccgg aagtgcttga
     2041 cattggggaa ttcagcgaga gcctgaccta ttgcatctcc cgccgtgcac agggtgtcac
     2101 gttgcaagac ctgcctgaaa ccgaactgcc cgctgttctg cagccggtcg cggaggccat
     2161 ggatgcgatc gctgcggccg atcttagcca gacgagcggg ttcggcccat tcggaccgca
     2221 aggaatcggt caatacacta catggcgtga tttcatatgc gcgattgctg atccccatgt
     2281 gtatcactgg caaactgtga tggacgacac cgtcagtgcg tccgtcgcgc aggctctcga
     2341 tgagctgatg ctttgggccg aggactgccc cgaagtccgg cacctcgtgc acgcggattt
     2401 cggctccaac aatgtcctga cggacaatgg ccgcataaca gcggtcattg actggagcga
     2461 ggcgatgttc ggggattccc aatacgaggt cgccaacatc ttcttctgga ggccgtggtt
     2521 ggcttgtatg gagcagcaga cgcgctactt cgagcggagg catccggagc ttgcaggatc
     2581 gccgcggctc cgggcgtata tgctccgcat tggtcttgac caactctatc agagcttggt
     2641 tgacggcaat ttcgatgatg cagcttgggc gcagggtcga tgcgacgcaa tcgtccgatc
     2701 cggagccggg actgtcgggc gtacacaaat cgcccgcaga agcgcggccg tctggaccga
     2761 tggctgtgta gaagtactcg ccgatagtgg aaaccgacgc cccagcactc gtccgagggc
     2821 aaaggaataa tcagtactga caataaaaag attcttgttt tcaagaactt gtcatttgta
     2881 tagttttttt atattgtagt tgttctattt taatcaaatg ttagcgtgat ttatattttt
     2941 tttcgcctcg acatcatctg cccagatgcg aagttaagtg cgcagaaagt aatatcatgc
     3001 gtcaatcgta tgtgaatgct ggtcgctata ctgctgtcga ttcgatacta acgccgccac
     3061 tagttctaga gcggccgcca ccgcggtgga gctccagctt ttgttccctt tagtgagggt
     3121 taattgcgcg cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
     3181 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     3241 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     3301 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     3361 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     3421 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     3481 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     3541 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     3601 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     3661 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     3721 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     3781 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     3841 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     3901 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     3961 ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc
     4021 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     4081 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     4141 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     4201 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     4261 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     4321 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     4381 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     4441 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     4501 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     4561 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     4621 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     4681 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     4741 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     4801 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     4861 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     4921 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     4981 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     5041 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     5101 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     5161 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     5221 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     5281 cccgaaaagt gccac
//
