LOCUS       sequence-487176-        5028 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV cloning vector for Flp dependent expression under the TRE 
            promoter. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5028)
  TITLE     Ritola Lab plasmids
REFERENCE   2  (bases 1 to 5028)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: TRE-FRT-MCSrev-WPRE-SV40 - GCH30.1
FEATURES             Location/Qualifiers
     source          1..5028
                     /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    1999..2032
                     /label=FRT3
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision with other FRT3 sites
                     (Bosch et al., 2015)"
     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     2133..2149
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2133..2149
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(2183..2199)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2184..2200)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     protein_bind    complement(2217..2250)
                     /label=FRT3
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision with other FRT3 sites
                     (Bosch et al., 2015)"
     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)."
     primer_bind     complement(2425..2449)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    join(2476..2494,2495..2517,2518..2536,2537..2559,2560..2578,
                     2579..2601,2602..2620,2621..2643,2644..2662,2663..2685,
                     2686..2704,2705..2727,2728..2746)
                     /label=tetracycline response element
                     /bound_moiety="tetracycline repressor TetR"
                     /note="contains seven copies of the tetracycline operator 
                     tetO"
     repeat_region   2777..2917
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2777..2906
                     /label=AAV2 ITR
     primer_bind     complement(2927..2944)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2927..2943)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2936..2958)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3156..3611
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3243..3262)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3453..3474
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3628..3647)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3747..3769
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3807..3825)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3893..3997
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3998..4066,4067..4858)
                     /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(4216..4235)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
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 ggaacttcga gctccaccgc ggtggcggcc gctctagaac tagtggatcc cccgggctgc
     2161 aggaattcga tatcaagctt atcgataccg tcgacctcga gggggggccc ggtaccgaag
     2221 ttcctatact atttgaagaa taggaacttc ggaataggaa cttcagagcg ctttggccgc
     2281 tccggccaaa gggaagttcc tatactttct agagaatagg aacttcggaa taggaacttc
     2341 accggtcaat tcggggccgc ggaggctgga tcggtcccgg tgtcttctat ggaggtcaaa
     2401 acagcgtgga tggcgtctcc aggcgatctg acggttcact aaacgagctc tgcttatata
     2461 ggtcgagttt accactccct atcagtgata gagaaaagtg aaagtcgagt ttaccactcc
     2521 ctatcagtga tagagaaaag tgaaagtcga gtttaccact ccctatcagt gatagagaaa
     2581 agtgaaagtc gagtttacca ctccctatca gtgatagaga aaagtgaaag tcgagtttac
     2641 cactccctat cagtgataga gaaaagtgaa agtcgagttt accactccct atcagtgata
     2701 gagaaaagtg aaagtcgagt ttaccactcc ctatcagtga tagagaaaag tgaaagtcga
     2761 gctcggtacc agatctagga acccctagtg atggagttgg ccactccctc tctgcgcgct
     2821 cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg
     2881 gcctcagtga gcgagcgagc gcgcagctgc ctgcaggcag cttggcactg gccgtcgttt
     2941 tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc
     3001 cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt
     3061 tgcgcagcct gaatggcgaa tggcgcctga tgcggtattt tctccttacg catctgtgcg
     3121 gtatttcaca ccgcatacgt caaagcaacc atagtacgcg ccctgtagcg gcgcattaag
     3181 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     3241 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     3301 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     3361 aaaacttgat ttgggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     3421 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     3481 actcaaccct atctcgggct attcttttga tttataaggg attttgccga tttcggccta
     3541 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac
     3601 gtttacaatt ttatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     3661 gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc
     3721 cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc
     3781 atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt
     3841 catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac
     3901 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     3961 ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt
     4021 cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct
     4081 ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga
     4141 tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag
     4201 cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca
     4261 actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga
     4321 aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag
     4381 tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc
     4441 ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa
     4501 tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt
     4561 gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg
     4621 gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt
     4681 tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg
     4741 gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat
     4801 ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact
     4861 gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa
     4921 aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt
     4981 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttc
//
