LOCUS       V035755                 9869 bp    DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   V035755
VERSION     V035755
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"
     promoter        2010..2213
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
                     /label="CMV promoter"
     CDS             2302..3018
                     /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"
     CDS             3055..4794
                     /label="METTL3(NM_019852)"
                     /note="METTL3(NM_019852)"
                     /gene="METTL3"
     promoter        4839..5050
                     /note="core&nbsp;promoter for human elongation factor
                     EF-1α"
                     /label="EF-1α core promoter"
     LTR             5063..5331
                     /note="truncated 5' long terminal repeat (LTR) from human
                     T-cell leukemia virus (HTLV) type 1"
                     /label="5' LTR (truncated)"
     CDS             5358..5957
                     /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    5964..6552
                     /note="woodchuck hepatitis virus&nbsp;posttranscriptional
                     regulatory element"
                     /label="WPRE"
     LTR             6626..6859
                     /note="self-inactivating&nbsp;3' long terminal repeat (LTR)
                     from HIV-1"
                     /label="3' LTR (ΔU3)"
     polyA_signal    6931..7052
                     /note="SV40 polyadenylation signal"
                     /label="SV40 poly(A) signal"
     rep_origin      7071..7206
                     /note="SV40 origin of replication"
                     /label="SV40 ori"
     primer_bind     complement(7244..7260)
                     /note="common sequencing primer, one of multiple similar
                     variants"
                     /label="M13 rev"
     protein_bind    7268..7284
                     /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(7292..7322)
                     /note="promoter for the E. coli lac operon"
                     /label="lac promoter"
     protein_bind    7337..7358
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
                     /label="CAP binding site"
     rep_origin      complement(7646..8234)
                     /direction=LEFT
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /label="ori"
     CDS             complement(8405..9265)
                     /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(9266..9370)
                     /gene="bla"
                     /label="AmpR promoter"
     primer_bind     9844..9860
                     /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 gtattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
     1981 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
     2041 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
     2101 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
     2161 tgacgcaaat gggcggtagg cgtgtacggt gggaggttta tataagcaga gctcgtttag
     2221 tgaaccgtca gatcgcctgg agacgccatc cacgctgttt tgacctccat agaagattct
     2281 agagctagcg aattcgccac catggtgagc aagggcgagg agctgttcac cggggtggtg
     2341 cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag
     2401 ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag
     2461 ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc
     2521 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac
     2581 gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg
     2641 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag
     2701 gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc
     2761 atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag
     2821 gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc
     2881 gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac
     2941 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc
     3001 atggacgagc tgtacaaggg cggcggaggg agtggaggag ggggaagtgg atccatgtcg
     3061 gacacgtgga gctctatcca ggcccacaag aagcagctgg actctctgcg ggagaggctg
     3121 cagcggaggc ggaagcagga ctcggggcac ttggatctac ggaatccaga ggcagcattg
     3181 tctccaacct tccgtagtga cagcccagtg cctactgcac ccacctctgg tggccctaag
     3241 cccagcacag cttcagcagt tcctgaatta gctacagatc ctgagttaga gaagaagttg
     3301 ctacaccacc tctctgatct ggccttaaca ttgcccactg atgctgtgtc catctgtctt
     3361 gccatctcca cgccagatgc tcctgccact caagatgggg tagaaagcct cctgcagaag
     3421 tttgcagctc aggagttgat tgaggtaaag cgaggtctcc tacaagatga tgcacatcct
     3481 actcttgtaa cctatgctga ccattccaag ctctctgcca tgatgggtgc tgtggcagaa
     3541 aagaagggcc ctggggaggt agcagggact gtcacagggc agaagcggcg tgcagaacag
     3601 gactcgacta cagtagctgc ctttgccagt tcgttagtct ctggtctgaa ctcttcagca
     3661 tcggaaccag caaaggagcc agccaagaaa tcaaggaaac atgctgcctc agatgttgat
     3721 ctggagatag agagccttct gaaccaacag tccactaagg aacaacagag caagaaggtc
     3781 agtcaggaga tcctagagct attaaatact acaacagcca aggaacaatc cattgttgaa
     3841 aaatttcgct ctcgaggtcg ggcccaagtg caagaattct gtgactatgg aaccaaggag
     3901 gagtgcatga aagccagtga tgctgatcga ccctgtcgca agctgcactt cagacgaatt
     3961 atcaataaac acactgatga gtctttaggt gactgctctt tccttaatac atgtttccac
     4021 atggatacct gcaagtatgt tcactatgaa attgatgctt gcatggattc tgaggcccct
     4081 ggcagcaaag accacacgcc aagccaggag cttgctctta cacagagtgt cggaggtgat
     4141 tccagtgcag accgactctt cccacctcag tggatctgtt gtgatatccg ctacctggac
     4201 gtcagtatct tgggcaagtt tgcagttgtg atggctgacc caccctggga tattcacatg
     4261 gaactgccct atgggaccct gacagatgat gagatgcgca ggctcaacat acccgtacta
     4321 caggatgatg gctttctctt cctctgggtc acaggcaggg ccatggagtt ggggagagaa
     4381 tgtctaaacc tctgggggta tgaacgggta gatgaaatta tttgggtgaa gacaaatcaa
     4441 ctgcaacgca tcattcggac aggccgtaca ggtcactggt tgaaccatgg gaaggaacac
     4501 tgcttggttg gtgtcaaagg aaatccccaa ggcttcaacc agggtctgga ttgtgatgtg
     4561 atcgtagctg aggttcgttc caccagtcat aaaccagatg aaatctatgg catgattgaa
     4621 agactatctc ctggcactcg caagattgag ttatttggac gaccacacaa tgtgcaaccc
     4681 aactggatca cccttggaaa ccaactggat gggatccacc tactagaccc agatgtggtt
     4741 gcacggttca agcaaaggta cccagatggt atcatctcta aacctaagaa tttataggcg
     4801 gccgcgaagg atctgcgatc gctccggtgc ccgtcagtgg gcagagcgca catcgcccac
     4861 agtccccgag aagttggggg gaggggtcgg caattgaacg ggtgcctaga gaaggtggcg
     4921 cggggtaaac tgggaaagtg atgtcgtgta ctggctccgc ctttttcccg agggtggggg
     4981 agaaccgtat ataagtgcag tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc
     5041 cagaacacag ctgaagcttc gaggggctcg catctctcct tcacgcgccc gccgccctac
     5101 ctgaggccgc catccacgcc ggttgagtcg cgttctgccg cctcccgcct gtggtgcctc
     5161 ctgaactgcg tccgccgtct aggtaagttt aaagctcagg tcgagaccgg gcctttgtcc
     5221 ggcgctccct tggagcctac ctagactcag ccggctctcc acgctttgcc tgaccctgct
     5281 tgctcaactc tacgtctttg tttcgttttc tgttctgcgc cgttacagat ccaagctgtg
     5341 accggcgcct acgctagatg accgagtaca agcccacggt gcgcctcgcc acccgcgacg
     5401 acgtcccccg ggccgtacgc accctcgccg ccgcgttcgc cgactacccc gccacgcgcc
     5461 acaccgtcga cccggaccgc cacatcgagc gggtcaccga gctgcaagaa ctcttcctca
     5521 cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg
     5581 tctggaccac gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca
     5641 tggccgagtt gagcggttcc cggctggccg cgcagcaaca gatggaaggc ctcctggcgc
     5701 cgcaccggcc caaggagccc gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc
     5761 agggcaaggg tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg
     5821 gggtgcccgc cttcctggag acctccgcgc cccgcaacct ccccttctac gagcggctcg
     5881 gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc
     5941 gcaagcccgg tgcctgagtc gacaatcaac ctctggatta caaaatttgt gaaagattga
     6001 ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct ttaatgcctt
     6061 tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat aaatcctggt
     6121 tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg
     6181 tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag ctcctttccg
     6241 ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc tgccttgccc
     6301 gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg tcggggaaat
     6361 catcgtcctt tccttggctg ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct
     6421 tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg
     6481 ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc tccctttggg
     6541 ccgcctcccc gcctggtacc tttaagacca atgacttaca aggcagctgt agatcttagc
     6601 cactttttaa aagaaaaggg gggactggaa gggctaattc actcccaacg aaaataagat
     6661 ctgctttttg cttgtactgg gtctctctgg ttagaccaga tctgagcctg ggagctctct
     6721 ggctaactag ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta
     6781 gtgtgtgccc gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca
     6841 gtgtggaaaa tctctagcag tagtagttca tgtcatctta ttattcagta tttataactt
     6901 gcaaagaaat gaatatcaga gagtgagagg aacttgttta ttgcagctta taatggttac
     6961 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt
     7021 tgtggtttgt ccaaactcat caatgtatct tatcatgtct ggctctagct atcccgcccc
     7081 taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg
     7141 cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga ggcttttttg
     7201 gaggcctaga cttttgcaga gacggcccaa attcgtaatc atggtcatag ctgtttcctg
     7261 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     7321 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     7381 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     7441 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     7501 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     7561 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     7621 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     7681 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     7741 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     7801 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     7861 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     7921 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     7981 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     8041 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta
     8101 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     8161 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     8221 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     8281 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     8341 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     8401 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     8461 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     8521 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     8581 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     8641 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     8701 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     8761 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     8821 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     8881 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     8941 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     9001 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     9061 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     9121 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     9181 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     9241 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     9301 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     9361 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca
     9421 tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg
     9481 atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag
     9541 cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg
     9601 gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat atgcggtgtg
     9661 aaataccgca cagatgcgta aggagaaaat accgcatcag gcgccattcg ccattcaggc
     9721 tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga
     9781 aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac
     9841 gttgtaaaac gacggccagt gccaagctg
//
