LOCUS       pBS-SKII-3XHA-fC        4653 bp DNA     circular SYN 20-AUG-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4653)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 14, 2022 from SnapGene 6.1.0
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4653
                     /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 benzylcytosine 
                     derivatives"
                     /label=CLIP-tag(TM)
                     /note="fast-reacting variant, optimized for mammalian 
                     expression"
                     /translation="MDKDCEMKRTTLDSPLGKLELSGCEQGLHRIIFLGKGTSAADAVE
                     VPAPAAVLGGPEPLIQATAWLNAYFHQPEAIEEFPVPALHHPVFQQESFTRQVLWKLLK
                     VVKFGEVISESHLAALVGNPAATAAVNTALDGNPVPILIPCHRVVQGDSDVGPYLGGLA
                     VKEWLLAHEGHRLGKPGLG"
     promoter        1458..1801
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1802..2193
                     /codon_start=1
                     /label=PhleoR/BleR
                     /translation="MGMTDQATPNLPSRDFDPTAAFYERLGFGIVFRDAGWMILQRGDL
                     MLEFFAHPGLDPLASWFSCCLRLDDLAEFYRQCKSVGIQETSSGYPRIHAPELQEWGGT
                     MAALVDPDGTLLRLIQNELLAGIS*S"
     terminator      2194..2391
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     promoter        complement(2465..2483)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2504..2520)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    2528..2544
                     /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(2552..2582)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2597..2618
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(2906..3494)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(3665..4525)
                     /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(4526..4630)
                     /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 gggctatccc 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 gatccaggcc accgcctggc tcaacgccta ctttcaccag cctgaggcca
     1081 tcgaggagtt ccctgtgcca gccctgcacc acccagtgtt ccagcaggag agctttaccc
     1141 gccaggtgct gtggaaactg ctgaaagtgg tgaagttcgg agaggtcatc agcgagagcc
     1201 acctggccgc cctggtgggc aatcccgccg ccaccgccgc cgtgaacacc gccctggacg
     1261 gaaatcccgt gcccattctg atcccctgcc accgggtggt gcagggcgac agcgacgtgg
     1321 ggccctacct 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 cgctgcagac ctgcgagcag ggaaacgctc ccctcacaga cgcgttgaat tgtccccacg
     1681 ccgcgcccct gtagagaaat ataaaaggtt aggatttgcc actgaggttc ttctttcata
     1741 tacttccttt taaaatcttg ctaggataca gttctcacat cacatccgaa cataaacaac
     1801 catgggtatg accgaccaag cgacgcccaa cctgccatca cgagatttcg atcccaccgc
     1861 cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct
     1921 ccagcgcggg gatctcatgc tggagttctt cgcccacccc gggctcgatc ccctcgcgag
     1981 ttggttcagc tgctgcctga ggctggacga cctcgcggag ttctaccggc agtgcaaatc
     2041 cgtcggcatc caggaaacca gcagcggcta tccgcgcatc catgcccccg aactgcagga
     2101 gtggggaggc acgatggccg ctttggtcga cccggacggg acgctcctgc gcctgataca
     2161 gaacgaattg cttgcaggca tctcatgatc agtactgaca ataaaaagat tcttgttttc
     2221 aagaacttgt catttgtata gtttttttat attgtagttg ttctatttta atcaaatgtt
     2281 agcgtgattt atattttttt tcgcctcgac atcatctgcc cagatgcgaa gttaagtgcg
     2341 cagaaagtaa tatcatgcgt caatcgtatg tgaatgctgg tcgctatact gctgtcgatt
     2401 cgatactaac gccgccacta gttctagagc ggccgccacc gcggtggagc tccagctttt
     2461 gttcccttta gtgagggtta attgcgcgct tggcgtaatc atggtcatag ctgtttcctg
     2521 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     2581 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     2641 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     2701 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     2761 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     2821 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     2881 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     2941 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     3001 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     3061 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     3121 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     3181 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     3241 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     3301 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta
     3361 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     3421 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     3481 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     3541 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     3601 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     3661 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     3721 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     3781 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     3841 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     3901 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     3961 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     4021 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     4081 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     4141 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     4201 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     4261 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     4321 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     4381 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     4441 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     4501 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     4561 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     4621 gggttccgcg cacatttccc cgaaaagtgc cac
//
