LOCUS       pBS-SKII-3XHA-eD        5226 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 5226)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 14, 2022 from SnapGene 6.1.0
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5226
                     /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..1334
                     /codon_start=1
                     /gene="folA"
                     /product="E. coli dihydrofolate reductase"
                     /label=ecDHFR
                     /translation="MISLIAALAVDRVIGMENAMPWNLPADLAWFKRNTLNKPVIMGRH
                     TWESIGRPLPGRKNIILSSQPGTDDRVTWVKSVDEAIAACGDVPEIMVIGGGRVYEQFL
                     PKAQKLYLTHIDAEVEGDTHFPDYEPDDWESVFSEFHDADAQNSHSYCFEILERR"
     gene            1389..2964
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     promoter        1389..1732
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1733..2761
                     /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      2767..2964
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     promoter        complement(3038..3056)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3077..3093)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3101..3117
                     /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(3125..3155)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3170..3191
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(3479..4067)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4238..5098)
                     /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(5099..5203)
                     /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 ttctatgatc agtctgattg cggcgttagc ggtagatcgc gttatcggca
      901 tggaaaacgc catgccgtgg aacctgcctg ccgatctcgc ctggtttaaa cgcaacacct
      961 taaataaacc cgtgattatg ggccgccata cctgggaatc aatcggtcgt ccgttgccag
     1021 gacgcaaaaa tattatcctc agcagtcaac cgggtacgga cgatcgcgta acgtgggtga
     1081 agtcggtgga tgaagccatc gcggcgtgtg gtgacgtacc agaaatcatg gtgattggcg
     1141 gcggtcgcgt ttatgaacag ttcttgccaa aagcgcaaaa actgtatctg acgcatatcg
     1201 acgcagaagt ggaaggcgac acccatttcc cggattacga gccggatgac tgggaatcgg
     1261 tattcagcga attccacgat gctgatgcgc agaactctca cagctattgc tttgagattc
     1321 tggagcggcg gtaatatata actgtctaga aataaagagt atcatctttc aaaggtcacc
     1381 cggccagcga catggaggcc cagaataccc tccttgacag tcttgacgtg cgcagctcag
     1441 gggcatgatg tgactgtcgc ccgtacattt agcccataca tccccatgta taatcatttg
     1501 catccataca ttttgatggc cgcacggcgc gaagcataaa ttacggctcc tcgctgcgga
     1561 cctgcgagca gggaaacgct cccctcacag acgcgttgaa ttgtccccac gccgcgcccc
     1621 tgtagagaaa tataaaaggt taggatttgc cactgaggtt cttctttcat atacttcctt
     1681 ttaaaatctt gctaggatac agttctcaca tcacatccga acataaacaa ccatgggtaa
     1741 aaagcctgaa ctcaccgcga cgtctgtcga gaagtttctg atcgaaaagt tcgacagcgt
     1801 ctccgacctg atgcagctct cggagggcga agaatctcgt gctttcagct tcgatgtagg
     1861 agggcgtgga tatgtcctgc gggtaaatag ctgcgccgat ggtttctaca aagatcgtta
     1921 tgtttatcgg cactttgcat cggccgcgct cccgattccg gaagtgcttg acattgggga
     1981 attcagcgag agcctgacct attgcatctc ccgccgtgca cagggtgtca cgttgcaaga
     2041 cctgcctgaa accgaactgc ccgctgttct gcagccggtc gcggaggcca tggatgcgat
     2101 cgctgcggcc gatcttagcc agacgagcgg gttcggccca ttcggaccgc aaggaatcgg
     2161 tcaatacact acatggcgtg atttcatatg cgcgattgct gatccccatg tgtatcactg
     2221 gcaaactgtg atggacgaca ccgtcagtgc gtccgtcgcg caggctctcg atgagctgat
     2281 gctttgggcc gaggactgcc ccgaagtccg gcacctcgtg cacgcggatt tcggctccaa
     2341 caatgtcctg acggacaatg gccgcataac agcggtcatt gactggagcg aggcgatgtt
     2401 cggggattcc caatacgagg tcgccaacat cttcttctgg aggccgtggt tggcttgtat
     2461 ggagcagcag acgcgctact tcgagcggag gcatccggag cttgcaggat cgccgcggct
     2521 ccgggcgtat atgctccgca ttggtcttga ccaactctat cagagcttgg ttgacggcaa
     2581 tttcgatgat gcagcttggg cgcagggtcg atgcgacgca atcgtccgat ccggagccgg
     2641 gactgtcggg cgtacacaaa tcgcccgcag aagcgcggcc gtctggaccg atggctgtgt
     2701 agaagtactc gccgatagtg gaaaccgacg ccccagcact cgtccgaggg caaaggaata
     2761 atcagtactg acaataaaaa gattcttgtt ttcaagaact tgtcatttgt atagtttttt
     2821 tatattgtag ttgttctatt ttaatcaaat gttagcgtga tttatatttt ttttcgcctc
     2881 gacatcatct gcccagatgc gaagttaagt gcgcagaaag taatatcatg cgtcaatcgt
     2941 atgtgaatgc tggtcgctat actgctgtcg attcgatact aacgccgcca ctagttctag
     3001 agcggccgcc accgcggtgg agctccagct tttgttccct ttagtgaggg ttaattgcgc
     3061 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
     3121 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct
     3181 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc
     3241 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt
     3301 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag
     3361 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca
     3421 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     3481 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     3541 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     3601 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     3661 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     3721 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     3781 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     3841 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     3901 actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct
     3961 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     4021 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     4081 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     4141 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     4201 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg
     4261 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt
     4321 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag
     4381 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc
     4441 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag
     4501 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca
     4561 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa
     4621 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga
     4681 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata
     4741 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca
     4801 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg
     4861 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg
     4921 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg
     4981 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag
     5041 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac
     5101 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca
     5161 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag
     5221 tgccac
//
