LOCUS       sequence-479187-        5285 bp    DNA     circular SYN 08-AUG-2025
DEFINITION  AAV cloning vector for Flp dependent expression under the GFAP 
            promoter. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5285)
  TITLE     Ritola Lab plasmids
REFERENCE   2  (bases 1 to 5285)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 8, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: GFAP-FRT-MCS-WPRE-SV40 - GCH33.1
FEATURES             Location/Qualifiers
     source          1..5285
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /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     956..978
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     975..991
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   1007..1147
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   1018..1147
                     /label=AAV2 ITR
     polyA_signal    1253..1374
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1290..1309)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1344..1363
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     misc_feature    1390..1978
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             1496..1507
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1905..1925
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    2061..2108
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     misc_feature    2109..2216
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     primer_bind     2125..2141
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     2126..2142
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2176..2192)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2176..2192)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    complement(2293..2340)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     repeat_region   3034..3174
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3034..3163
                     /label=AAV2 ITR
     primer_bind     complement(3184..3201)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3184..3200)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3193..3215)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3413..3868
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3500..3519)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3710..3731
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3885..3904)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4004..4026
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4064..4082)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4150..4254
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4255..4323,4324..5115)
                     /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"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga ccatgattac gaattgcctg caggcagctg
     1021 cgcgctcgct cgctcactga ggccgcccgg gcaaagcccg ggcgtcgggc gacctttggt
     1081 cgcccggcct cagtgagcga gcgagcgcgc agagagggag tggccaactc catcactagg
     1141 ggttcctatc gatggcgcgc ctaccacatt tgtagaggtt ttacttgctt taaaaaacct
     1201 cccacatctc cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt
     1261 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     1321 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     1381 ctgtgtacag cggggaggcg gcccaaaggg agatccgact cgtctgaggg cgaaggcgaa
     1441 gacgcggaag aggccgcaga gccggcagca ggccgcggga aggaaggtcc gctggattga
     1501 gggccgaagg gacgtagcag aaggacgtcc cgcgcagaat ccaggtggca acacaggcga
     1561 gcagccaagg aaaggacgat gatttccccg acaacaccac ggaattgtca gtgcccaaca
     1621 gccgagcccc tgtccagcag cgggcaaggc aggcggcgat gagttccgcc gtggcaatag
     1681 ggagggggaa agcgaaagtc ccggaaagga gctgacaggt ggtggcaatg ccccaaccag
     1741 tgggggttgc gtcagcaaac acagtgcaca ccacgccacg ttgcctgaca acgggccaca
     1801 actcctcata aagagacagc aaccaggatt tatacaagga ggagaaaatg aaagccatac
     1861 gggaagcaat agcatgatac aaaggcatta aagcagcgta tccacatagc gtaaaaggag
     1921 caacatagtt aagaatacca gtcaatcttt cacaaatttt gtaatccaga ggttgattgc
     1981 tagcgaagtt cctattccga agttcctatt cttcaaatag tataggaact tcttggatca
     2041 aaataggagc gatcgcaatg gaagttccta ttccgaagtt cctattctct agaaagtata
     2101 ggaacttcgg taccgggccc cccctcgagg tcgacggtat cgataagctt gatatcgaat
     2161 tcctgcagcc cgggggatcc actagttcta gagcggccgc caccgcggtg gagctcgaag
     2221 ttcctatact atttgaagaa taggaacttc ggaataggaa cttcagagcg ctttggccgc
     2281 tccggccaaa gggaagttcc tatactttct agagaatagg aacttcggaa taggaacttc
     2341 accggtgcga gcagcggagg tgatgcgtct cctctccaac ctgctctggc tctgctcgct
     2401 cctgggatgc gagggcttta tgaaggagtg ggctagactg gcgatgcccg ggtgcccctg
     2461 gcaacacccc ctggcatagc ctgagggggt ggggttgggc cgcagcccag ctatggggag
     2521 agctctcccc tcacccattt gtgtccttgg acttctggga agcagacagc cctcctctcc
     2581 ccccctgcca cccactgtcc atgcctgggc atcccctgat cccctttcct gggcacaggc
     2641 tgaatagagc cttgttctcc accgccggtg gcgccacaca gcaagggcag cccctagggg
     2701 gctgtgctgc ttttatccca agatgccagg ctgtcaggct ggggtgcaat gcaagccccc
     2761 tgctcaatgg gcttctcgga aggcatttga ggggggctgg ggtgcggcgg gaagcaggca
     2821 ctgtgcccac cccgaacatt gtgtctgtgc caaggtgagt cattcactgg gcatgaagag
     2881 gaggccctgg ggggcggcca gcccaatgct gccaggtctg cttgcagaca aggcctggct
     2941 gtctgcctcc tccccagctc acagagccag ctcaggcccc cgagcctctg tcagagcagc
     3001 gtcccctact ccacaccagg atatgttaga tctaggaacc cctagtgatg gagttggcca
     3061 ctccctctct gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc gcccgacgcc
     3121 cgggctttgc ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg caggcagctt
     3181 ggcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa
     3241 tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga
     3301 tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg cgcctgatgc ggtattttct
     3361 ccttacgcat ctgtgcggta tttcacaccg catacgtcaa agcaaccata gtacgcgccc
     3421 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     3481 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     3541 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     3601 cggcacctcg accccaaaaa acttgatttg ggtgatggtt cacgtagtgg gccatcgccc
     3661 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     3721 ttccaaactg gaacaacact caaccctatc tcgggctatt cttttgattt ataagggatt
     3781 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     3841 tttaacaaaa tattaacgtt tacaatttta tggtgcactc tcagtacaat ctgctctgat
     3901 gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct
     3961 tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt
     4021 cagaggtttt caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta
     4081 tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg
     4141 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     4201 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
     4261 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
     4321 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
     4381 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
     4441 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
     4501 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     4561 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     4621 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     4681 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     4741 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
     4801 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     4861 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     4921 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     4981 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     5041 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     5101 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     5161 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     5221 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     5281 tcttc
//
