LOCUS       V029181                 8908 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V029181
VERSION     V029181
KEYWORDS    .
SOURCE      .
  ORGANISM  .
            .
FEATURES             Location/Qualifiers
     LTR             234..414
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
                     /label="5' LTR (truncated)"
     misc_feature    458..583
                     /note="packaging signal of human immunodeficiency virus
                     type 1"
                     /label="HIV-1 Ψ"
     misc_feature    1076..1309
                     /note="The Rev response element (RRE) of HIV-1 allows for
                     Rev-dependent mRNA export from the nucleus to the
                     cytoplasm."
                     /label="RRE"
     CDS             1493..1537
                     /codon_start=1
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /product="antigenic peptide corresponding to amino acids
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik
                     et al., 2013)"
                     /transl_table=1
                     /translation="KNEQELLELDKWASL"
                     /label="gp41 peptide"
     misc_feature    1804..1921
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
                     /label="cPPT/CTS"
     promoter        2011..2214
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /label="CMV promoter"
     CDS             2317..2934
                     /label="Twist1(NM_011658)"
                     /note="Twist1(NM_011658)"
                     /gene="Twist1"
     CDS             2941..3006
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     promoter        3050..3261
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             3274..3542
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     regulatory      3571..3580
                     /note="vertebrate consensus sequence for strong initiation
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
                     /label="Kozak sequence"
     CDS             3601..4263
                     /codon_start=1
                     /product="green fluorescent protein 2 from Pontellina
                     plumata, also known as ppluGFP2 (Shagin et al., 2004)"
                     /transl_table=1
                     /translation="PAMEIECRITGTLNGVEFELVGGGEGTPKQGRMTNKMKSTKGALT
                     FSPYLLSHVMGYGFYHFGTYPSGYENPFLHAINNGGYTNTRIEKYEDGGVLHVSFSYRY
                     EAGRVIGDFKVVGTGFPEDSVIFTDKIIRSNATVEHLHPMGDNVLVGSFARTFSLRDGG
                     YYSFVVDSHMHFKSAIHPSILQNGGPMFAFRRVEELHSNTELGIVEYQHAFKTPIAFA"
                     /label="CopGFP"
     CDS             4333..4386
                     /codon_start=1
                     /note="Eukaryotic ribosomes fail to insert a peptide bond
                     between the Gly and Pro residues, yielding separate
                     polypeptides."
                     /product="2A peptide from&nbsp;Thosea asigna virus capsid
                     protein"
                     /protein_id=""
                     /transl_table=1
                     /translation="EGRGSLLTCGDVEENPGP"
                     /label="T2A"
     CDS             4387..4986
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /note="confers resistance to puromycin"
                     /product="puromycin N-acetyltransferase"
                     /transl_table=1
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA*"
                     /label="PuroR"
     misc_feature    4987..5575
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             5649..5882
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    5954..6075
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6115..6250
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6283..6299)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6307..6323
                     /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-β-D-thiogalactopyranoside (IPTG)."
                     /label="lac operator"
     promoter        complement(6331..6361)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6376..6397
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6685..7273)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7444..8304)
                     /codon_start=1
                     /gene="bla"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /product="β-lactamase"
                     /transl_table=1
                     /translation="MSIQHFRVALIPFFAAFCLPVFA,HPETLVKVKDAEDQLGARVGY
                     IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEY
                     SPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDR
                     WEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRS
                     ALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGA
                     SLIKHW*"
                     /label="AmpR"
     promoter        complement(8305..8409)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     8883..8899
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 fwd"
ORIGIN
        1 acgcgtgtag tcttatgcaa tactcttgta gtcttgcaac atggtaacga tgagttagca
       61 acatgcctta caaggagaga aaaagcaccg tgcatgccga ttggtggaag taaggtggta
      121 cgatcgtgcc ttattaggaa ggcaacagac gggtctgaca tggattggac gaaccactga
      181 attgccgcat tgcagagata ttgtatttaa gtgcctagct cgatacaata aacgggtctc
      241 tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac ccactgctta
      301 agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact
      361 ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct agcagtggcg
      421 cccgaacagg gacctgaaag cgaaagggaa accagagctc tctcgacgca ggactcggct
      481 tgctgaagcg cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc caaaaatttt
      541 gactagcgga ggctagaagg agagagatgg gtgcgagagc gtcagtatta agcgggggag
      601 aattagatcg cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa aatataaatt
      661 aaaacatata gtatgggcaa gcagggagct agaacgattc gcagttaatc ctggcctgtt
      721 agaaacatca gaaggctgta gacaaatact gggacagcta caaccatccc ttcagacagg
      781 atcagaagaa cttagatcat tatataatac agtagcaacc ctctattgtg tgcatcaaag
      841 gatagagata aaagacacca aggaagcttt agacaagata gaggaagagc aaaacaaaag
      901 taagaccacc gcacagcaag cggccactga tcttcagacc tggaggagga gatatgaggg
      961 acaattggag aagtgaatta tataaatata aagtagtaaa aattgaacca ttaggagtag
     1021 cacccaccaa ggcaaagaga agagtggtgc agagagaaaa aagagcagtg ggaataggag
     1081 ctttgttcct tgggttcttg ggagcagcag gaagcactat gggcgcagcc tcaatgacgc
     1141 tgacggtaca ggccagacaa ttattgtctg gtatagtgca gcagcagaac aatttgctga
     1201 gggctattga ggcgcaacag catctgttgc aactcacagt ctggggcatc aagcagctcc
     1261 aggcaagaat cctggctgtg gaaagatacc taaaggatca acagctcctg gggatttggg
     1321 gttgctctgg aaaactcatt tgcaccactg ctgtgccttg gaatgctagt tggagtaata
     1381 aatctctgga acagattgga atcacacgac ctggatggag tgggacagag aaattaacaa
     1441 ttacacaagc ttaatacact ccttaattga agaatcgcaa aaccagcaag aaaagaatga
     1501 acaagaatta ttggaattag ataaatgggc aagtttgtgg aattggttta acataacaaa
     1561 ttggctgtgg tatataaaat tattcataat gatagtagga ggcttggtag gtttaagaat
     1621 agtttttgct gtactttcta tagtgaatag agttaggcag ggatattcac cattatcgtt
     1681 tcagacccac ctcccaaccc cgaggggacc cgacaggccc gaaggaatag aagaagaagg
     1741 tggagagaga gacagagaca gatccattcg attagtgaac ggatctcgac ggtatcggtt
     1801 aacttttaaa agaaaagggg ggattggggg gtacagtgca ggggaaagaa tagtagacat
     1861 aatagcaaca gacatacaaa ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt
     1921 tatcgatact agtattatgc ccagtacatg accttatggg actttcctac ttggcagtac
     1981 atctacgtat tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg
     2041 cgtggatagc ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg
     2101 agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca
     2161 ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggttt atataagcag agctcgttta
     2221 gtgaaccgtc agatcgcctg gagacgccat ccacgctgtt ttgacctcca tagaagattc
     2281 tagagctagc ctgatacgaa ctcggaattc gccaccatga tgcaggacgt gtccagctcg
     2341 ccagtctctc cggccgacga cagcctgagc aacagcgagg aggagccgga ccggcagcag
     2401 ccggcgagcg gcaagcgcgg ggctcgcaag agacgcagca gtcggcgcag cgcgggcggc
     2461 agcgcggggc ccggcggggc cacgggcggg ggcatcggag gcggcgacga gccaggcagc
     2521 ccggcccagg gcaagcgcgg caagaaatct gcgggcggag gcggcggcgg cggcgcgggc
     2581 ggaggtggtg gcggcggcgg cggcagcagc agcgggggcg ggagcccgca gtcgtacgag
     2641 gagctgcaga cccagcgggt catggctaac gtgcgggagc gccagcgcac gcagtcgctg
     2701 aacgaggcgt tcgccgccct gcgcaagatc atccccacgc tgccctcgga caagctgagc
     2761 aagattcaga ccctcaaact ggcggccagg tacatcgact tcctgtacca ggtcctgcag
     2821 agcgacgagc tggactccaa gatggcaagc tgcagctatg tggcccacga gcggctcagc
     2881 tacgccttct ccgtctggag gatggagggg gcctggtcca tgtccgcgtc ccacggatcc
     2941 gactacaaag accatgacgg tgattataaa gatcatgaca tcgactacaa ggatgacgat
     3001 gacaagtagc ggccgcgaag gatctgcgat cgctccggtg cccgtcagtg ggcagagcgc
     3061 acatcgccca cagtccccga gaagttgggg ggaggggtcg gcaattgaac gggtgcctag
     3121 agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt actggctccg cctttttccc
     3181 gagggtgggg gagaaccgta tataagtgca gtagtcgccg tgaacgttct ttttcgcaac
     3241 gggtttgccg ccagaacaca gctgaagctt cgaggggctc gcatctctcc ttcacgcgcc
     3301 cgccgcccta cctgaggccg ccatccacgc cggttgagtc gcgttctgcc gcctcccgcc
     3361 tgtggtgcct cctgaactgc gtccgccgtc taggtaagtt taaagctcag gtcgagaccg
     3421 ggcctttgtc cggcgctccc ttggagccta cctagactca gccggctctc cacgctttgc
     3481 ctgaccctgc ttgctcaact ctacgtcttt gtttcgtttt ctgttctgcg ccgttacaga
     3541 tccaagctgt gaccggcgcc tacgctagac gccaccatgg agagcgacga gagcggcctg
     3601 cccgccatgg agatcgagtg ccgcatcacc ggcaccctga acggcgtgga gttcgagctg
     3661 gtgggcggcg gagagggcac ccccaagcag ggccgcatga ccaacaagat gaagagcacc
     3721 aaaggcgccc tgaccttcag cccctacctg ctgagccacg tgatgggcta cggcttctac
     3781 cacttcggca cctaccccag cggctacgag aaccccttcc tgcacgccat caacaacggc
     3841 ggctacacca acacccgcat cgagaagtac gaggacggcg gcgtgctgca cgtgagcttc
     3901 agctaccgct acgaggccgg ccgcgtgatc ggcgacttca aggtggtggg caccggcttc
     3961 cccgaggaca gcgtgatctt caccgacaag atcatccgca gcaacgccac cgtggagcac
     4021 ctgcacccca tgggcgataa cgtgctggtg ggcagcttcg cccgcacctt cagcctgcgc
     4081 gacggcggct actacagctt cgtggtggac agccacatgc acttcaagag cgccatccac
     4141 cccagcatcc tgcagaacgg gggccccatg ttcgccttcc gccgcgtgga ggagctgcac
     4201 agcaacaccg agctgggcat cgtggagtac cagcacgcct tcaagacccc catcgccttc
     4261 gccagatccc gcgctcagtc gtccaattct gccgtggacg gcaccgccgg acccggctcc
     4321 accggatctc gcgagggcag aggaagtctt ctaacatgcg gtgacgtgga ggagaatccc
     4381 ggccctatga ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga cgtccccagg
     4441 gccgtacgca ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat
     4501 ccggaccgcc acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac gcgcgtcggg
     4561 ctcgacatcg gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt ctggaccacg
     4621 ccggagagcg tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat ggccgagttg
     4681 agcggttccc ggctggccgc gcagcaacag atggaaggcc tcctggcgcc gcaccggccc
     4741 aaggagcccg cgtggttcct ggccaccgtc ggcgtctcgc ccgaccacca gggcaagggt
     4801 ctgggcagcg ccgtcgtgct ccccggagtg gaggcggccg agcgcgccgg ggtgcccgcc
     4861 ttcctggaga cctccgcgcc ccgcaacctc cccttctacg agcggctcgg cttcaccgtc
     4921 accgccgacg tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg caagcccggt
     4981 gcctgaaatc aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat
     5041 gttgctcctt ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct
     5101 tcccgtatgg ctttcatttt ctcctccttg tataaatcct ggttgctgtc tctttatgag
     5161 gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc
     5221 cccactggtt ggggcattgc caccacctgt cagctccttt ccgggacttt cgctttcccc
     5281 ctccctattg ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct
     5341 cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga aatcatcgtc ctttccttgg
     5401 ctgctcgcct gtgttgccac ctggattctg cgcgggacgt ccttctgcta cgtcccttcg
     5461 gccctcaatc cagcggacct tccttcccgc ggcctgctgc cggctctgcg gcctcttccg
     5521 cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt gggccgcctc cccgcctggt
     5581 acctttaaga ccaatgactt acaaggcagc tgtagatctt agccactttt taaaagaaaa
     5641 ggggggactg gaagggctaa ttcactccca acgaagataa gatctgcttt ttgcttgtac
     5701 tgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc
     5761 actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt
     5821 gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag
     5881 cagtagtagt tcatgtcatc ttattattca gtatttataa cttgcaaaga aatgaatatc
     5941 agagagtgag aggaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     6001 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     6061 catcaatgta tcttatcatg tctggctcta gctatcccgc ccctaactcc gcccatcccg
     6121 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     6181 tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg aggaggcttt
     6241 tttggaggcc tagacttttg cagagaccaa attcgtaatc atgtcatagc tgtttcctgt
     6301 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     6361 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     6421 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     6481 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     6541 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     6601 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     6661 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     6721 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     6781 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     6841 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     6901 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     6961 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     7021 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     7081 tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat
     7141 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     7201 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     7261 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     7321 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     7381 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     7441 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     7501 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     7561 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     7621 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     7681 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     7741 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     7801 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     7861 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     7921 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     7981 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     8041 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     8101 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     8161 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     8221 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     8281 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     8341 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     8401 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat
     8461 gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtctcgcgc gtttcggtga
     8521 tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc
     8581 ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg
     8641 ctggcttaac tatgcggcat cagagcagat tgtactgaga gtgcaccata tgcggtgtga
     8701 aataccgcac agatgcgtaa ggagaaaata ccgcatcagg cgccattcgc cattcaggct
     8761 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
     8821 agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg
     8881 ttgtaaaacg acggccagtg ccaagctg
//
