LOCUS       V044825                 9118 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V044825
VERSION     V044825
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..1920
                     /note="central polypurine tract and central termination
                     sequence of HIV-1"
                     /label="cPPT/CTS"
     enhancer        1933..2236
                     /note="human cytomegalovirus immediate early enhancer"
                     /label="CMV enhancer"
     promoter        2237..2440
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /label="CMV promoter"
     CDS             2475..4337
                     /label="KLHL40(NM_152393)"
                     /note="KLHL40(NM_152393)"
                     /gene="KLHL40"
     CDS             4344..4409
                     /codon_start=1
                     /product="three tandem FLAG® epitope tags, followed by an
                     enterokinase cleavage site"
                     /transl_table=1
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
                     /label="3xFLAG"
     CDS             4419..4475
                     /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 porcine teschovirus-1
                     polyprotein"
                     /transl_table=1
                     /translation="ATNFSLLKQAGDVEENPGP"
                     /label="P2A"
     CDS             4488..5207
                     /codon_start=1
                     /note="mammalian codon-optimized"
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /transl_table=1
                     /translation="M,V,SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKL
                     TLKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFK
                     DDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNG
                     IKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMV
                     LLEFVTAAGITLGMDELYK*"
                     /label="EGFP"
     misc_feature    5214..5801
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             5875..6108
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6180..6301
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      6320..6455
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(6493..6509)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    6517..6533
                     /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(6541..6571)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    6586..6607
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(6895..7483)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(7654..8514)
                     /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(8515..8619)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     9093..9109
                     /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 attacaaaat tcaaaatttt
     1921 atcgatacta gtcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
     1981 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
     2041 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
     2101 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
     2161 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
     2221 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
     2281 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
     2341 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
     2401 cggtaggcgt gtacggtggg aggtctatat aagcagagct ggtttagtga accgtcagat
     2461 ccgctagcgc caccatggcg ctgggcttgg agcaggcgga ggagcagcgg ttgtaccagc
     2521 agacgctcct gcaagacggg ctcaaagaca tgctggacca tggcaagttc ctcgactgtg
     2581 tggtgcgggc gggcgagcgc gagttcccgt gccatcgcct ggtgctggcc gcctgcagcc
     2641 cctacttccg ggcgcgcttt ctagccgagc cggagcgcgc gggcgagctg cacctggagg
     2701 aggtgtcccc ggacgtggtg gcccaggtgc tgcactacct gtacacatca gagatcgcgc
     2761 tggatgaggc gagcgtgcag gatttgttcg ccgcggcaca ccgcttccag atcccttcca
     2821 tcttcaccat ctgcgtgtcc ttcctgcaga agcgcctgtg cctctccaac tgcttggccg
     2881 tcttccgtct cggcctcctg ctcgactgcg cgcgtctcgc cgtggctgcc cgcgacttca
     2941 tctgcgctca cttcacgctg gtggcgcgcg acgctgactt cctcggactc tcggccgacg
     3001 agctcatcgc catcatctcc agcgacggcc ttaacgtgga gaaggaggag gcagtgttcg
     3061 aggcggtgat gcggtgggcg ggtagcggcg acgccgaggc gcaggctgag cgccagcgcg
     3121 cgctgcccac cgtcttcgag agcgtgcgct gccgcttgct gccgcgcgcc tttctggaaa
     3181 gccgcgtgga gcgccaccct ctcgtgcgtg cccagcccga gttgctgcgc aaggtgcaga
     3241 tggtgaagga tgcacacgag ggccgcatca ccacgctgcg gaagaaaaag aaggggaagg
     3301 atggagccgg ggccaaggag gctgataagg gcacaagcaa agccaaagca gaggaggatg
     3361 aggaggccga acgtatcctt cctgggatcc tcaatgacac cctgcgcttc ggcatgttcc
     3421 tgcaggatct catcttcatg atcagtgagg agggcgctgt ggcctacgat ccagcagcca
     3481 acgagtgcta ctgtgcttcc ctctccaacc aggtccccaa gaaccacgtc agcctggtta
     3541 ccaaggagaa ccaggtcttc gtggctggag gcctcttcta caacgaagac aacaaagagg
     3601 accccatgag cgcatacttc ctgcagtttg accatctgga ctcagagtgg ctggggatgc
     3661 caccgctgcc ctcgccccgc tgcctctttg gcctgggaga agctctcaac tccatctacg
     3721 tggtcggtgg cagagagatc aaggacggcg agcgctgcct ggactcggtc atgtgctacg
     3781 acaggctgtc attcaaatgg ggtgaatcgg acccgctgcc ttacgtggtg tatggccaca
     3841 cagtgctctc ccacatggac cttgtctacg taattggcgg caaaggcagt gacaggaagt
     3901 gcctgaacaa gatgtgcgtc tatgacccca agaagtttga gtggaaggag ctggcaccca
     3961 tgcagaccgc ccgctcactc tttggggcca ctgtccatga tggccgcatt atcgtggcag
     4021 ctggggtcac cgacacaggg ctgaccagtt ctgccgaagt gtacagcatc acagacaaca
     4081 agtgggcacc cttcgaggcc ttcccacagg agcgtagctc actcagcctg gtcagcctgg
     4141 tgggtaccct ctatgccatt ggtggctttg ccacactgga gacggagtct ggagagctgg
     4201 ttcccacaga gctcaatgac atctggaggt ataacgagga ggagaagaaa tgggagggtg
     4261 tcctgcggga gatcgcctat gcagcaggtg ccaccttcct accagtgcgg ctcaatgtgc
     4321 tgtgcctgac taagatggtc gacgactaca aagaccatga cggtgattat aaagatcatg
     4381 acatcgacta caaggatgac gatgacaagg gaagcggagc cactaacttc tccctgttga
     4441 aacaagcagg ggatgtcgaa gagaatcccg ggccagaatt cgccaccatg gtgagcaagg
     4501 gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg
     4561 gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc
     4621 tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc
     4681 tgacctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag cacgacttct
     4741 tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg
     4801 gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg
     4861 agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca
     4921 actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc atcaaggtga
     4981 acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac cactaccagc
     5041 agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagcaccc
     5101 agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg
     5161 tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagtaagga tccaatcaac
     5221 ctctggatta caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta
     5281 cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt
     5341 tcattttctc ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg
     5401 ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg
     5461 gcattgccac cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca
     5521 cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca
     5581 ctgacaattc cgtggtgttg tcggggaagc tgacgtcctt tccatggctg ctcgcctgtg
     5641 ttgccacctg gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag
     5701 cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt ctccgccttc
     5761 gccctcagac gagtcggatc tccctttggc cgcctccccg cctggtacct ttaagaccaa
     5821 tgacttacaa ggcagctgta gatcttagcc actttttaaa agaaaagggg ggactggaag
     5881 ggctaattca ctcccaacga aaataagatc tgctttttgc ttgtactggg tctctctggt
     5941 tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc
     6001 aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta
     6061 actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt agtagttcat
     6121 gtcatcttat tattcagtat ttataacttg caaagaaatg aatatcagag agtgagagga
     6181 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
     6241 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
     6301 atcatgtctg gctctagcta tcccgcccct aactccgccc agttccgccc attctccgcc
     6361 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct
     6421 attccagaag tagtgaggag gcttttttgg aggcctagac ttttgcagag acggcccaaa
     6481 ttcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     6541 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     6601 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     6661 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     6721 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
     6781 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     6841 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     6901 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     6961 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     7021 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     7081 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     7141 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     7201 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     7261 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     7321 cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     7381 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     7441 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     7501 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     7561 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     7621 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     7681 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     7741 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     7801 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     7861 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     7921 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     7981 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     8041 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     8101 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     8161 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     8221 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     8281 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     8341 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     8401 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     8461 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     8521 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     8581 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     8641 acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac
     8701 gaggcccttt cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct
     8761 cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg
     8821 cgcgtcagcg ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat cagagcagat
     8881 tgtactgaga gtgcaccata tgcggtgtga aataccgcac agatgcgtaa ggagaaaata
     8941 ccgcatcagg cgccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg
     9001 ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg
     9061 ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg ccaagctg
//
