LOCUS       62056_3430        6737 bp DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6737)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6737
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(1..105)
                     /label=AmpR promoter
     rep_origin      complement(131..586)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     primer_bind     728..744
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        1442..1645
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     CDS             1731..1748
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     CDS             1764..1787
                     /codon_start=1
                     /label=Strep-Tag II
                     /note="peptide that binds Strep-Tactin(R), an engineered 
                     form
                     of streptavidin"
                     /translation="WSHPQFEK"
     CDS             1797..1850
                     /codon_start=1
                     /label=T2A
                     /note="2A peptide from Thosea asigna virus capsid protein"
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             1872..1895
                     /codon_start=1
                     /label=FLAG
                     /note="FLAG(R) epitope tag, followed by an enterokinase
                     cleavage site"
                     /translation="DYKDDDDK"
     promoter        1939..2150
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     LTR             2163..2431
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
     CDS             2475..2501
                     /codon_start=1
                     /label=HA
                     /note="HA (human influenza hemagglutinin) epitope tag"
                     /translation="YPYDVPDYA"
     CDS             2508..2537
                     /codon_start=1
                     /label=Myc
                     /note="Myc (human c-Myc proto-oncogene) epitope tag"
                     /translation="EQKLISEEDL"
     CDS             2547..2600
                     /codon_start=1
                     /label=T2A
                     /note="2A peptide from Thosea asigna virus capsid protein"
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             2601..3197
                     /codon_start=1
                     /label=PuroR
                     /note="puromycin N-acetyltransferase"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     polyA_signal    3308..3440
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4716..4732)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(4740..4756)
                     /label=lac operator
                     /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(4764..4794)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(4809..4830)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(5118..5706)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(5880..6737)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
ORIGIN
        1 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
       61 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
      121 agtgccacct aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
      181 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
      241 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
      301 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
      361 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
      421 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
      481 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
      541 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc
      601 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg
      661 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca
      721 cgacgttgta aaacgacggc cagtgagcgc gcctcgttca ttcacgtttt tgaacccgtg
      781 gaggacgggc agactcgcgg tgcaaatgtg ttttacagcg tgatggagca gatgaagatg
      841 ctcgacacgc tgcagaacac gcagctagat taaccctaga aagataatca tattgtgacg
      901 tacgttaaag ataatcatgc gtaaaattga cgcatgtgtt ttatcggtct gtatatcgag
      961 gtttatttat taatttgaat agatattaag ttttattata tttacactta catactaata
     1021 ataaattcaa caaacaattt atttatgttt atttatttat taaaaaaaaa caaaaactca
     1081 aaatttcttc tataaagtaa caaaactttt atgagggaca gccccccccc aaagccccca
     1141 gggatgtaat tacgtccctc ccccgctagg gggcagcagc gagccgcccg gggctccgct
     1201 ccggtccggc gctccccccg catccccgag ccggcagcgt gcggggacag cccgggcacg
     1261 gggaaggtgg cacgggatcg ctttcctctg aacgcttctc gctgctcttt gagcctgcag
     1321 acacctgggg ggatacgggg aaaaggcctc caaggcctac tagtattatg cccagtacat
     1381 gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat
     1441 ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt
     1501 tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga
     1561 ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg
     1621 gtgggaggtc tatataagca gagctcgttt agtgaaccgt cagatcgcct ggagacgcca
     1681 tccacgctgt tttgacctcc atagaagatt ctagagccac catggaattc catcaccatc
     1741 accatcacgt cgacggcgga tcctggagcc acccgcagtt cgagaaagga agcggagagg
     1801 gcagaggaag tctgctaaca tgcggtgacg tcgaggagaa tcctggacct atgctcgaga
     1861 ccggtagatc tgattacaag gatgacgacg ataagtgagc ggccgcaagg atctgcgatc
     1921 gctccggtgc ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg
     1981 gaggggtcgg caattgaacg ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg
     2041 atgtcgtgta ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag
     2101 tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc cagaacacag ctgaagcttc
     2161 gaggggctcg catctctcct tcacgcgccc gccgccctac ctgaggccgc catccacgcc
     2221 ggttgagtcg cgttctgccg cctcccgcct gtggtgcctc ctgaactgcg tccgccgtct
     2281 aggtaagttt aaagctcagg tcgagaccgg gcctttgtcc ggcgctccct tggagcctac
     2341 ctagactcag ccggctctcc acgctttgcc tgaccctgct tgctcaactc tacgtctttg
     2401 tttcgttttc tgttctgcgc cgttacagat ccaagctgtg accggcgcct acgctagacg
     2461 ccaccatggc tagctaccca tacgacgtcc cagactacgc tggtaccgag cagaaactca
     2521 tctctgaaga ggatctggga agcggagagg gcagaggaag tcttctaaca tgcggtgacg
     2581 tggaggagaa tcccggccct atgaccgagt acaagcccac ggtgcgcctc gccacccgcg
     2641 acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc
     2701 gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa gaactcttcc
     2761 tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg
     2821 cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag atcggcccgc
     2881 gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa ggcctcctgg
     2941 cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc
     3001 accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg gccgagcgcg
     3061 ccggggtgcc cgccttcctg gagacctccg cgccccgcaa cctccccttc tacgagcggc
     3121 tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga
     3181 cccgcaagcc cggtgcctga aatcaacctc tggattacaa aatttgtgaa agattgactg
     3241 gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt
     3301 atcagttaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     3361 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     3421 tgtatcttat catgtctgga attgactcaa atgatgtcaa ttagtctatc agaagctatc
     3481 tggtctccct tccgggggac aagacatccc tgtttaatat ttaaacagca gtgttcccaa
     3541 actgggttct tatatccctt gctctggtca accaggttgc agggtttcct gtcctcacag
     3601 gaacgaagtc cctaaagaaa cagtggcagc caggtttagc cccggaattg actggattcc
     3661 ttttttaggg cccattggta tggctttttc cccgtatccc cccaggtgtc tgcaggctca
     3721 aagagcagcg agaagcgttc agaggaaagc gatcccgtgc caccttcccc gtgcccgggc
     3781 tgtccccgca cgctgccggc tcggggatgc ggggggagcg ccggaccgga gcggagcccc
     3841 gggcggctcg ctgctgcccc ctagcggggg agggacgtaa ttacatccct gggggctttg
     3901 ggggggggct gtccctgata tctataacaa gaaaatatat atataataag ttatcacgta
     3961 agtagaacat gaaataacaa tataattatc gtatgagtta aatcttaaaa gtcacgtaaa
     4021 agataatcat gcgtcatttt gactcacgcg gtcgttatag ttcaaaatca gtgacactta
     4081 ccgcattgac aagcacgcct cacgggagct ccaagcggcg actgagatgt cctaaatgca
     4141 cagcgacgga ttcgcgctat ttagaaagag agagcaatat ttcaagaatg catgcgtcaa
     4201 ttttacgcag actatctttc tagggttaat ctagctgcat caggatcata tcgtcgggtc
     4261 ttttttccgg ctcagtcatc gcccaagctg gcgctatctg ggcatcgggg aggaagaagc
     4321 ccgtgccttt tcccgcgagg ttgaagcggc atggaaagag tttgccgagg atgactgctg
     4381 ctgcattgac gttgagcgaa aacgcacgtt taccatgatg attcgggaag gtgtggccat
     4441 gcacgccttt aacggtgaac tgttcgttca ggccacctgg gataccagtt cgtcgcggct
     4501 tttccggaca cagttccgga tggtcagccc gaagcgcatc agcaacccga acaataccgg
     4561 cgacagccgg aactgccgtg ccggtgtgca gattaatgac agcggtgcgg cgctgggata
     4621 ttacgtcagc gaggacgggt atcctggctg gatgccgcag aaatggacat ggataccccg
     4681 tgagttaccc ggcgggcgcg cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat
     4741 tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg
     4801 ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag
     4861 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     4921 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     4981 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     5041 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     5101 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     5161 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     5221 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     5281 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     5341 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     5401 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     5461 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     5521 ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct
     5581 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     5641 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     5701 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
     5761 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat
     5821 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac
     5881 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt
     5941 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt
     6001 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag
     6061 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct
     6121 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt
     6181 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc
     6241 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt
     6301 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg
     6361 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg
     6421 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct
     6481 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc
     6541 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt
     6601 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt
     6661 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg
     6721 aaatgttgaa tactcat
//
