LOCUS       pAAV_EfIa_DIO_NC        7613 bp DNA     circular SYN 02-MAY-2023
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 7613)
  AUTHORS   Hochgeschwender
  TITLE     Direct Submission
  JOURNAL   Exported Mar 4, 2024 from SnapGene 7.1.0
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7613
                     /mol_type="other DNA"
                     /organism="recombinant plasmid"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     primer_bind     complement(23..34)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     84..95
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        242..1420
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          473..1411
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1368..1388
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     primer_bind     1368..1388
                     /label=C63
     protein_bind    1451..1484
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites."
     protein_bind    1535..1568
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     CDS             complement(1586..2578)
                     /label=ChRmine
     CDS             complement(2580..2623)
                     /codon_start=1
                     /label=15aa linker
                     /translation="GDPLVQCGGIAGSA"
     CDS             complement(join(2630..3126,3127..3132,3133..3808))
                     /codon_start=1
                     /label=NCS3
                     /translation="VSKGEEDNMASLPATHELHIFGSINGVDFDMVGQGTGNPNDGYEE
                     LNLKSTKGDLQFSPWILVPHIGYGFHQYLPYPDGMSPFQAAMVDGSGYQVHRTMQFEDG
                     ASLTVNYRYTYEGSHIKGEAQVKGTGFPADGPVMTNSLTAADWCRSKKTYPNDKTIIST
                     FKWSYTTGNGKRYRSTARTTYTFAKPMAANYLKNQPMYVFRKTELKHSKTELNFKEWQK
                     AFTGFEDFVGDWEQIAAYNLDQVLEQGGVSSVLQTLAVSVTPIQRIVRSGENGLKIDIH
                     VIIPYEGLSADQMAHIEEVFKVVYPVDDHHFKVIMEYGTLVIDGVTPNMLNYFGRPYEG
                     IAVFDGNKITVTGTLWNGNKIIDERLITPEGSMLFRVTINGVTGYRLLKKISN"
     primer_bind     complement(2671..2690)
                     /label=C650
     primer_bind     complement(3330..3349)
                     /label=C644
     primer_bind     3330..3349
                     /label=C646
     primer_bind     complement(3791..3808)
                     /label=Lifeact-F
     primer_bind     complement(3791..3808)
                     /label=C564
     CDS             complement(3809..3859)
                     /codon_start=1
                     /label=GLuc signal sequence
                     /translation="MGVKVLFALICIAVAEA"
     regulatory      3856..3865
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(3890..3923)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites."
     protein_bind    complement(3974..4007)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    4032..4620
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4085..4105)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     primer_bind     complement(4099..4119)
                     /label=D527
     CDS             complement(4503..4514)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4653..4764
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4876..5016
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4876..5005
                     /label=AAV2 ITR
     primer_bind     complement(4922..4933)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     4983..4994
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      5091..5546
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5178..5197)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5388..5409
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5563..5582)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5682..5704
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5742..5760)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5828..5932
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5933..6001,6002..6793)
                     /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"
     primer_bind     complement(6151..6170)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6964..7552
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7432..7451
                     /label=C398
     primer_bind     7453..7472
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7502..7521
                     /label=C397
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtaagct ttgcaaagat ggataaagtt
      181 ttaaacagag aggaatcttt gcagctaatg gaccttctag gtcttgaaag gagtgggaat
      241 tggctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg agaagttggg
      301 gggaggggtc ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa actgggaaag
      361 tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc
      421 agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac aggtaagtgc
      481 cgtgtgtggt tcccgcgggc ctggcctctt tacgggttat ggcccttgcg tgccttgaat
      541 tacttccacc tggctgcagt acgtgattct tgatcccgag cttcgggttg gaagtgggtg
      601 ggagagttcg aggccttgcg cttaaggagc cccttcgcct cgtgcttgag ttgaggcctg
      661 gcctgggcgc tggggccgcc gcgtgcgaat ctggtggcac cttcgcgcct gtctcgctgc
      721 tttcgataag tctctagcca tttaaaattt ttgatgacct gctgcgacgc tttttttctg
      781 gcaagatagt cttgtaaatg cgggccaaga tctgcacact ggtatttcgg tttttggggc
      841 cgcgggcggc gacggggccc gtgcgtccca gcgcacatgt tcggcgaggc ggggcctgcg
      901 agcgcggcca ccgagaatcg gacgggggta gtctcaagct ggccggcctg ctctggtgcc
      961 tggcctcgcg ccgccgtgta tcgccccgcc ctgggcggca aggctggccc ggtcggcacc
     1021 agttgcgtga gcggaaagat ggccgcttcc cggccctgct gcagggagct caaaatggag
     1081 gacgcggcgc tcgggagagc gggcgggtga gtcacccaca caaaggaaaa gggcctttcc
     1141 gtcctcagcc gtcgcttcat gtgactccac ggagtaccgg gcgccgtcca ggcacctcga
     1201 ttagttctcg agcttttgga gtacgtcgtc tttaggttgg ggggaggggt tttatgcgat
     1261 ggagtttccc cacactgagt gggtggagac tgaagttagg ccagcttggc acttgatgta
     1321 attctccttg gaatttgccc tttttgagtt tggatcttgg ttcattctca agcctcagac
     1381 agtggttcaa agtttttttc ttccatttca ggtgtcgtga ggtaccggat cctctagagt
     1441 cgactccgga ataacttcgt ataggatact ttatacgaag ttatgcagaa tggtagctgg
     1501 attgtagctg ctattagcaa tatgaaacct cttaataact tcgtatagca tacattatac
     1561 gaagttatgg cgcgccacgc gttcacacgt tgatgtcgat ctggtccagg gggatgtact
     1621 cgccctcgct ggtgatcctg ctcttggcgg ccgcagacag tctccgcagg cgctcgccat
     1681 tgctgctcag cctcatatcc agctccagct tctggaactc atcctcgtcg tgtcccagct
     1741 gctccagcag ccgaaacagc tcctcatcat aggtctttct gaaccgggcc agggcaaagc
     1801 cgtaacagga cttggccacg atgtcgcaca cgcagtgggc cacctctgtc cagttatcgt
     1861 cgatccagcc gaagcctcta gggcagatgg cccacaccag gggaaacaga atccacagca
     1921 tggagaaggt aaacacggcg aacttcagca cgtgcaggct gtgctcggcg cctgtatcct
     1981 gggccagctg ggccagccgg ctatactgct tggccacgat gctcatcacg atagagtagg
     2041 cgaagataaa ccagaaacag ccaaagccat accaggcgta ggcgccattc ctcagcctag
     2101 ggtcgccctc tgcgtagaac tcggccagca cgccagacat caggatggca aagatgatgg
     2161 cgctacaaga cacgcgatag ggggccctca gctgatacag cagatcgtac accagcattg
     2221 ggcaggtcag catgtagtcg gcatacctgt aattgccgaa gccgatgatg aaggggccca
     2281 catcgccgaa gttgtggccc agtgtgtcgg ccacgatctt gccgaagtag ctcagcatgg
     2341 caaagaaatt cacaaacagg gccaggttct gctcctcctg atagccggcg taaaaggagg
     2401 ttctcttgct agggccccga gatgtccagg actcatagcc caggtagaat gtggtgattg
     2461 tcagcacgat aaagcaagac cagtgggcgc cgatggccac ggagttcagc ttggtatcca
     2521 gataccagcc gtccattgtg cccacgtaga acatctggtc ggtgcctggt gcgtgtgcgg
     2581 tggcggatcc ggcaattcca ccacactgga ctagtgggtc gccctcgagg ttagagatct
     2641 tcttcagcag acggtaaccg gtaacaccgt tgatggtaac acggaacagc atagaacctt
     2701 caggggtgat cagacgttcg tcgatgatct tgttaccgtt ccacagggta ccggtaacgg
     2761 tgatcttatt accgtcgaaa acagcgatac cttcgtaagg acgaccgaag tagttcagca
     2821 tgttaggggt aacaccgtcg ataaccaggg taccgtattc catgataacc ttgaaatgat
     2881 ggtcgtcaac agggtaaaca accttgaaaa cttcttcgat atgagccatc tggtcagcag
     2941 acagaccttc gtaagggatg ataacatgga tgtcgatctt cagaccgttt tcaccagaac
     3001 gaacgatacg ctggataggg gtaacagaaa cagccagggt ctgcagaaca gaagaaacac
     3061 caccctgttc cagaacctgg tccaggttgt aagcagcgat ctgttcccag tcaccaacga
     3121 agtcttcaaa tccggtaaag gccttttgcc actccttgaa gttgagctcg gtcttggagt
     3181 gcttgagctc cgtcttacgg aacacgtaca tcggctggtt cttcagatag ttagccgcca
     3241 ttggcttggc aaaggtgtag gtggtccgcg cagtgctccg gtagcgcttg ccatttccag
     3301 tggtgtaact ccacttaaag gtactgatga tggttttgtc gttggggtaa gtcttcttcg
     3361 acctgcacca gtccgcagcg gtcagcgagt tggtcatcac aggaccgtca gcagggaaac
     3421 cagtcccctt cacctgggcc tctcctttga tgtggcttcc ctcgtaggtg tagcggtagt
     3481 taacagtaag ggaggcacca tcttcaaact gcattgtgcg atggacttgg tatccggagc
     3541 catctaccat ggcggcctgg aaaggcgaca tcccgtcagg gtagggcagg tactgatgga
     3601 agccataccc gatatgaggg accagaatcc agggggagaa ctggaggtca cccttggtgg
     3661 acttcaggtt taactcctca taaccatcat ttggattgcc ggtgccctga cccaccatgt
     3721 caaagtccac accgttgatg gagccaaaga tgtgtaactc atgtgtcgct gggaggctag
     3781 ccatgttatc ctcctcgccc ttgctcacgg cctcggccac agcgatgcag atcagggcaa
     3841 acagaacttt gactcccatg gtggcggatc caagcttacc ggtgctagca taacttcgta
     3901 taaagtatcc tatacgaagt tatttgcctt aacccagaaa ttatcactgt tattctttag
     3961 aatggtgcaa agaataactt cgtataatgt atgctatacg aagttatgaa ttcgatatca
     4021 agcttatcga taatcaacct ctggattaca aaatttgtga aagattgact ggtattctta
     4081 actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta
     4141 ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt
     4201 atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg
     4261 caacccccac tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt
     4321 tccccctccc tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag
     4381 gggctcggct gttgggcact gacaattccg tggtgttgtc ggggaaatca tcgtcctttc
     4441 cttggctgct cgcctatgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc
     4501 cttcggccct caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc
     4561 ttccgcgtct tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc
     4621 atcgataccg agcgctgctc gagagatctc gactgtgcct tctagttgcc agccatctgt
     4681 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     4741 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     4801 tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc atgcacgtgc
     4861 ggaccgagcg gccgcaggaa cccctagtga tggagttggc cactccctct ctgcgcgctc
     4921 gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg
     4981 cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg tattttctcc
     5041 ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag caaccatagt acgcgccctg
     5101 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     5161 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     5221 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     5281 gcacctcgac cccaaaaaac ttgatttggg tgatggttca cgtagtgggc catcgccctg
     5341 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     5401 ccaaactgga acaacactca accctatctc gggctattct tttgatttat aagggatttt
     5461 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt
     5521 taacaaaata ttaacgttta caattttatg gtgcactctc agtacaatct gctctgatgc
     5581 cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg
     5641 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca
     5701 gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt
     5761 tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg
     5821 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     5881 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
     5941 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     6001 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
     6061 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     6121 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     6181 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     6241 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     6301 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
     6361 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     6421 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     6481 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
     6541 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     6601 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     6661 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     6721 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     6781 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
     6841 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     6901 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     6961 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     7021 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     7081 cttcagcaga gcgcagatac caaatactgt ccttctagtg tagccgtagt taggccacca
     7141 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     7201 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     7261 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     7321 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     7381 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     7441 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     7501 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     7561 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgt
//
