LOCUS       Exported                7867 bp DNA     circular SYN 22-AUG-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pSIH1-H1-copGFP-T2A-Puro
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7867)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..7867
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        6..232
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             233..413
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    460..585
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1078..1311
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1496..1540
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    1807..1923
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2013..2216
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     regulatory      2288..2297
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2318..2980
                     /codon_start=1
                     /product="green fluorescent protein 2 from Pontellina 
                     plumata, also known as ppluGFP2 (Shagin et al., 2004)"
                     /label=CopGFP
                     /translation="PAMEIECRITGTLNGVEFELVGGGEGTPKQGRMTNKMKSTKGALT
                     FSPYLLSHVMGYGFYHFGTYPSGYENPFLHAINNGGYTNTRIEKYEDGGVLHVSFSYRY
                     EAGRVIGDFKVVGTGFPEDSVIFTDKIIRSNATVEHLHPMGDNVLVGSFARTFSLRDGG
                     YYSFVVDSHMHFKSAIHPSILQNGGPMFAFRRVEELHSNTELGIVEYQHAFKTPIAFA"
     CDS             3050..3103
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             3104..3703
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     misc_feature    3704..4292
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     promoter        4404..4618
                     /label=H1 promoter
                     /note="human H1 RNA promoter"
     LTR             4661..4841
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     polyA_signal    4913..5034
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      5074..5209
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5242..5258)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    5266..5282
                     /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(5290..5320)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5335..5356
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(5644..6232)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(6403..7263)
                     /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(7264..7368)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7842..7858
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
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 cgatacataa acgggtctct
      241 ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc cactgcttaa
      301 gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt tgtgtgactc
      361 tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta gcagtggcgc
      421 ccgaacaggg acttgaaagc gaaagggaaa ccagaggagc tctctcgacg caggactcgg
      481 cttgctgaag cgcgcacggc aagaggcgag gggcggcgac tggtgagtac gccaaaaatt
      541 ttgactagcg gaggctagaa ggagagagat gggtgcgaga gcgtcagtat taagcggggg
      601 agaattagat cgcgatggga aaaaattcgg ttaaggccag ggggaaagaa aaaatataaa
      661 ttaaaacata tagtatgggc aagcagggag ctagaacgat tcgcagttaa tcctggcctg
      721 ttagaaacat cagaaggctg tagacaaata ctgggacagc tacaaccatc ccttcagaca
      781 ggatcagaag aacttagatc attatataat acagtagcaa ccctctattg tgtgcatcaa
      841 aggatagaga taaaagacac caaggaagct ttagacaaga tagaggaaga gcaaaacaaa
      901 agtaagacca ccgcacagca agcggccgct gatcttcaga cctggaggag gagatatgag
      961 ggacaattgg agaagtgaat tatataaata taaagtagta aaaattgaac cattaggagt
     1021 agcacccacc aaggcaaaga gaagagtggt gcagagagaa aaaagagcag tgggaatagg
     1081 agctttgttc cttgggttct tgggagcagc aggaagcact atgggcgcag cgtcaatgac
     1141 gctgacggta caggccagac aattattgtc tggtatagtg cagcagcaga acaatttgct
     1201 gagggctatt gaggcgcaac agcatctgtt gcaactcaca gtctggggca tcaagcagct
     1261 ccaggcaaga atcctggctg tggaaagata cctaaaggat caacagctcc tggggatttg
     1321 gggttgctct ggaaaactca tttgcaccac tgctgtgcct tggaatgcta gttggagtaa
     1381 taaatctctg gaacagattt ggaatcacac gacctggatg gagtgggaca gagaaattaa
     1441 caattacaca agcttaatac actccttaat tgaagaatcg caaaaccagc aagaaaagaa
     1501 tgaacaagaa ttattggaat tagataaatg ggcaagtttg tggaattggt ttaacataac
     1561 aaattggctg tggtatataa aattattcat aatgatagta ggaggcttgg taggtttaag
     1621 aatagttttt gctgtacttt ctatagtgaa tagagttagg cagggatatt caccattatc
     1681 gtttcagacc cacctcccaa ccccgagggg acccgacagg cccgaaggaa tagaagaaga
     1741 aggtggagag agagacagag acagatccat tcgattagtg aacggatctc gacggtatcg
     1801 gttaactttt aaaagaaaag gggggattgg ggggtacagt gcaggggaaa gaatagtaga
     1861 cataatagca acagacatac aaactaaaga attacaaaaa caaattacaa aattcaaaat
     1921 tttatcgata ctagtattat gcccagtaca tgaccttatg ggactttcct acttggcagt
     1981 acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg
     2041 ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg
     2101 ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc
     2161 cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ttatataagc agagctcgtt
     2221 tagtgaaccg tcagatcgcc tggagacgcc atccacgctg ttttgacctc catagaagat
     2281 tctagacgcc accatggaga gcgacgagag cggcctgccc gccatggaga tcgagtgccg
     2341 catcaccggc accctgaacg gcgtggagtt cgagctggtg ggcggcggag agggcacccc
     2401 caagcagggc cgcatgacca acaagatgaa gagcaccaaa ggcgccctga ccttcagccc
     2461 ctacctgctg agccacgtga tgggctacgg cttctaccac ttcggcacct accccagcgg
     2521 ctacgagaac cccttcctgc acgccatcaa caacggcggc tacaccaaca cccgcatcga
     2581 gaagtacgag gacggcggcg tgctgcacgt gagcttcagc taccgctacg aggccggccg
     2641 cgtgatcggc gacttcaagg tggtgggcac cggcttcccc gaggacagcg tgatcttcac
     2701 cgacaagatc atccgcagca acgccaccgt ggagcacctg caccccatgg gcgataacgt
     2761 gctggtgggc agcttcgccc gcaccttcag cctgcgcgac ggcggctact acagcttcgt
     2821 ggtggacagc cacatgcact tcaagagcgc catccacccc agcatcctgc agaacggggg
     2881 ccccatgttc gccttccgcc gcgtggagga gctgcacagc aacaccgagc tgggcatcgt
     2941 ggagtaccag cacgccttca agacccccat cgccttcgcc agatcccgcg ctcagtcgtc
     3001 caattctgcc gtggacggca ccgccggacc cggctccacc ggatctcgcg agggcagagg
     3061 aagtcttcta acatgcggtg acgtggagga gaatcccggc cctatgaccg agtacaagcc
     3121 cacggtgcgc ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc
     3181 gttcgccgac taccccgcca cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt
     3241 caccgagctg caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt
     3301 cgcggacgac ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc
     3361 ggtgttcgcc gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca
     3421 gcaacagatg gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc
     3481 caccgtcggc gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc
     3541 cggagtggag gcggccgagc gcgccggggt gcccgccttc ctggagacct ccgcgccccg
     3601 caacctcccc ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga
     3661 aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc tgaaatcaac ctctggatta
     3721 caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg
     3781 atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc
     3841 ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca
     3901 acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac
     3961 cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact
     4021 catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc
     4081 cgtggtgttg tcggggaaat catcgtcctt tccttggctg ctcgcctgtg ttgccacctg
     4141 gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc
     4201 ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt ctccgccttc gccctcagac
     4261 gagtcggatc tccctttggg ccgcctcccc gcctggtacc tttaagacca atgacttaca
     4321 aggcagctgt agatcttagc cactttttaa aagaaaaggg gggactggaa gggctaattc
     4381 actcccaacg atgtcaagaa ttggaacgct gacgtcatca acccgctcca aggaatcgcg
     4441 ggcccagtgt cactaggcgg gaacacccag cgcgcgtgcg ccctggcagg aagatggctg
     4501 tgagggacag gggagtggcg ccctgcaata tttgcatgtc gctatgtgtt ctgggaaatc
     4561 accataaacg tgaaatgtct ttggatttgg gaatcttata agttctgtat gagaccactt
     4621 ggatcctctg aattcttcga ttctgctttt tgcttctact gggtctctct ggttagacca
     4681 gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag
     4741 cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag
     4801 atccctcaga ccctttcagt cagtgtggaa aatctctagc agtagtagtt catgtcatct
     4861 tattattcag tatttataac ttgcaaagaa atgaatatca gagagtgaga ggaacttgtt
     4921 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     4981 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     5041 ctggctctag ctatcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     5101 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     5161 tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct agacttttgc
     5221 agagaccaaa ttcgtaatca tgtcatagct gtttcctgtg tgaaattgtt atccgctcac
     5281 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     5341 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     5401 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     5461 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     5521 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     5581 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     5641 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     5701 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     5761 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     5821 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     5881 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     5941 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     6001 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     6061 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     6121 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     6181 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     6241 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     6301 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     6361 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     6421 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     6481 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     6541 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     6601 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     6661 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     6721 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     6781 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     6841 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     6901 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     6961 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     7021 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     7081 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     7141 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     7201 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     7261 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     7321 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     7381 aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct ataaaaatag
     7441 gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat gacggtgaaa acctctgaca
     7501 catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc
     7561 ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc tggcttaact atgcggcatc
     7621 agagcagatt gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag
     7681 gagaaaatac cgcatcaggc gccattcgcc attcaggctg cgcaactgtt gggaagggcg
     7741 atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg
     7801 attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggccagtgc
     7861 caagctg
//
