LOCUS       Exported                8734 bp ds-DNA     circular SYN 12-JAN-2024
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pLV-Puro-CMV-CuO_MCS-T2A-mCherry
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8734)
  AUTHORS   mrk006
  TITLE     Direct Submission
  JOURNAL   Exported Mar 17, 2025 from SnapGene Viewer 4.0.2
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8734
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        3..229
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             230..410
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    457..582
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1075..1308
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1493..1537
                     /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    1803..1920
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     protein_bind    1938..1958
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     enhancer        1959..2262
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2263..2466
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     protein_bind    2467..2494
                     /label=CuO
                     /bound_moiety="CymR"
                     /note="CymR-binding P2 operator sequence from the p-cmt 
                     operon of Pseudomonas putida (Mullick et al., 2006)"
     protein_bind    2494..2518
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    2525..2632
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     primer_bind     2542..2558
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2592..2608)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             2642..2695
                     /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             2696..3406
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     protein_bind    complement(3407..3431)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    3449..4037
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             complement(3920..3931)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     promoter        4068..4567
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     CDS             4584..5183
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     LTR             5255..5488
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    5560..5681
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      5721..5856
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        complement(5877..5895)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5905..5921)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      6063..6518
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        6544..6648
                     /gene="bla"
                     /label=AmpR promoter
     CDS             6649..7509
                     /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"
     rep_origin      7680..8268
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    8556..8577
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        8592..8622
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8630..8646
                     /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)."
     primer_bind     8654..8670
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        8691..8709
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 aatgtagtct tatgcaatac tcttgtagtc ttgcaacatg gtaacgatga gttagcaaca
       61 tgccttacaa ggagagaaaa agcaccgtgc atgccgattg gtggaagtaa ggtggtacga
      121 tcgtgcctta ttaggaaggc aacagacggg tctgacatgg attggacgaa ccactgaatt
      181 gccgcattgc agagatattg tatttaagtg cctagctcga tacataaacg ggtctctctg
      241 gttagaccag atctgagcct gggagctctc tggctaacta gggaacccac tgcttaagcc
      301 tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg
      361 taactagaga tccctcagac ccttttagtc agtgtggaaa atctctagca gtggcgcccg
      421 aacagggact tgaaagcgaa agggaaacca gaggagctct ctcgacgcag gactcggctt
      481 gctgaagcgc gcacggcaag aggcgagggg cggcgactgg tgagtacgcc aaaaattttg
      541 actagcggag gctagaagga gagagatggg tgcgagagcg tcagtattaa gcgggggaga
      601 attagatcgc gatgggaaaa aattcggtta aggccagggg gaaagaaaaa atataaatta
      661 aaacatatag tatgggcaag cagggagcta gaacgattcg cagttaatcc tggcctgtta
      721 gaaacatcag aaggctgtag acaaatactg ggacagctac aaccatccct tcagacagga
      781 tcagaagaac ttagatcatt atataataca gtagcaaccc tctattgtgt gcatcaaagg
      841 atagagataa aagacaccaa ggaagcttta gacaagatag aggaagagca aaacaaaagt
      901 aagaccaccg cacagcaagc ggccgctgat cttcagacct ggaggaggag atatgaggga
      961 caattggaga agtgaattat ataaatataa agtagtaaaa attgaaccat taggagtagc
     1021 acccaccaag gcaaagagaa gagtggtgca gagagaaaaa agagcagtgg gaataggagc
     1081 tttgttcctt gggttcttgg gagcagcagg aagcactatg ggcgcagcgt caatgacgct
     1141 gacggtacag gccagacaat tattgtctgg tatagtgcag cagcagaaca atttgctgag
     1201 ggctattgag gcgcaacagc atctgttgca actcacagtc tggggcatca agcagctcca
     1261 ggcaagaatc ctggctgtgg aaagatacct aaaggatcaa cagctcctgg ggatttgggg
     1321 ttgctctgga aaactcattt gcaccactgc tgtgccttgg aatgctagtt ggagtaataa
     1381 atctctggaa cagatttgga 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 ggtatcgcta
     1801 gcttttaaaa gaaaaggggg gattgggggg tacagtgcag gggaaagaat agtagacata
     1861 atagcaacag acatacaaac taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt
     1921 actagtgatt atcggatcaa ctttgtatag aaaagttgcg ttacataact tacggtaaat
     1981 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     2041 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     2101 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
     2161 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
     2221 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
     2281 tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
     2341 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac
     2401 aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
     2461 agagctaaca aacagacaat ctggtctgtt tgtacaagtt tgtacaaaaa agcaggctgc
     2521 caccggtacc gggccccccc tcgaggtcga cggtatcgat aagcttgata tcgaattcct
     2581 gcagcccggg ggatccacta gttctagagc ggccgccacc gcggtggagc tcggaagcgg
     2641 agagggcagg ggaagtcttc taacatgcgg ggacgtggag gaaaatcccg gccccatggt
     2701 gagcaagggc gaggaggata acatggccat catcaaggag ttcatgcgct tcaaggtgca
     2761 catggagggc tccgtgaacg gccacgagtt cgagatcgag ggcgagggcg agggccgccc
     2821 ctacgagggc acccagaccg ccaagctgaa ggtgaccaag ggtggccccc tgcccttcgc
     2881 ctgggacatc ctgtcccctc agttcatgta cggctccaag gcctacgtga agcaccccgc
     2941 cgacatcccc gactacttga agctgtcctt ccccgagggc ttcaagtggg agcgcgtgat
     3001 gaacttcgag gacggcggcg tggtgaccgt gacccaggac tcctccctgc aggacggcga
     3061 gttcatctac aaggtgaagc tgcgcggcac caacttcccc tccgacggcc ccgtaatgca
     3121 gaagaagacc atgggctggg aggcctcctc cgagcggatg taccccgagg acggcgccct
     3181 gaagggcgag atcaagcaga ggctgaagct gaaggacggc ggccactacg acgctgaggt
     3241 caagaccacc tacaaggcca agaagcccgt gcagctgccc ggcgcctaca acgtcaacat
     3301 caagttggac atcacctccc acaacgagga ctacaccatc gtggaacagt acgaacgcgc
     3361 cgagggccgc cactccaccg gcggcatgga cgagctgtac aagtaaaccc agctttcttg
     3421 tacaaagtgg tgataatcga attccgataa tcaacctctg gattacaaaa tttgtgaaag
     3481 attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat
     3541 gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc
     3601 ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg
     3661 cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct
     3721 ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct
     3781 tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg
     3841 gaagctgacg tcctttccat ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac
     3901 gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct
     3961 gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct
     4021 ttgggccgcc tccccgcatc gggaattccc gcggttcgaa ttctaccggg taggggaggc
     4081 gcttttccca aggcagtctg gagcatgcgc tttagcagcc ccgctgggca cttggcgcta
     4141 cacaagtggc ctctggcctc gcacacattc cacatccacc ggtaggcgcc aaccggctcc
     4201 gttctttggt ggccccttcg cgccaccttc tactcctccc ctagtcagga agttcccccc
     4261 cgccccgcag ctcgcgtcgt gcaggacgtg acaaatggaa gtagcacgtc tcactagtct
     4321 cgtgcagatg gacagcaccg ctgagcaatg gaagcgggta ggcctttggg gcagcggcca
     4381 atagcagctt tgctccttcg ctttctgggc tcagaggctg ggaaggggtg ggtccggggg
     4441 cgggctcagg ggcgggctca ggggcggggc gggcgcccga aggtcctccg gaggcccggc
     4501 attctgcacg cttcaaaagc gcacgtctgc cgcgctgttc tcctcttcct catctccggg
     4561 cctttcgacc tcacgtggcc accatgaccg agtacaagcc cacggtgcgc ctcgccaccc
     4621 gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac taccccgcca
     4681 cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg caagaactct
     4741 tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac ggcgccgcgg
     4801 tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc gagatcggcc
     4861 cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg gaaggcctcc
     4921 tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcggc gtctcgcccg
     4981 accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag gcggccgagc
     5041 gcgccggggt gcccgccttc ctggagacct ccgcgccccg caacctcccc ttctacgagc
     5101 ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc acctggtgca
     5161 tgacccgcaa gcccggtgcc tgaggtacct ttaagaccaa tgacttacaa ggcagctgta
     5221 gatcttagcc actttttaaa agaaaagggg ggactggaag ggctaattca ctcccaacga
     5281 agacaagatc tgctttttgc ttgtactggg tctctctggt tagaccagat ctgagcctgg
     5341 gagctctctg gctaactagg gaacccactg cttaagcctc aataaagctt gccttgagtg
     5401 cttcaagtag tgtgtgcccg tctgttgtgt gactctggta actagagatc cctcagaccc
     5461 ttttagtcag tgtggaaaat ctctagcagt agtagttcat gtcatcttat tattcagtat
     5521 ttataacttg caaagaaatg aatatcagag agtgagagga acttgtttat tgcagcttat
     5581 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     5641 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg gctctagcta
     5701 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
     5761 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc
     5821 tattccagaa gtagtgagga ggcttttttg gaggcctagg gacgtaccca attcgcccta
     5881 tagtgagtcg tattacgcgc gctcactggc cgtcgtttta caacgtcgtg actgggaaaa
     5941 ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa
     6001 tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg
     6061 ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     6121 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     6181 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     6241 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg
     6301 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     6361 tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt
     6421 ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt
     6481 taacgcgaat tttaacaaaa tattaacgct tacaatttag gtggcacttt tcggggaaat
     6541 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     6601 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
     6661 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
     6721 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
     6781 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
     6841 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
     6901 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
     6961 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
     7021 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     7081 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     7141 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg
     7201 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     7261 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     7321 gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt
     7381 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     7441 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     7501 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
     7561 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     7621 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     7681 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     7741 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     7801 agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc
     7861 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     7921 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     7981 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     8041 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaagag
     8101 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     8161 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     8221 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     8281 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg
     8341 ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc
     8401 cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcccaata
     8461 cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt
     8521 cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct cactcattag
     8581 gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga
     8641 taacaatttc acacaggaaa cagctatgac catgattacg ccaagcgcgc aattaaccct
     8701 cactaaaggg aacaaaagct ggagctgcaa gctt
//
