LOCUS       sequence-173897-        7294 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway destination transfer plasmid for making AAV. Contains a DIO 
            cassette for cre-induction. With cre, insert will flip backwards, 
            causing it to turn off (cre-off).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7294)
  AUTHORS   Cammack AJ, Moudgil A, Chen J, Vasek MJ, Shabsovich M, McCullough K,
            Yen A, Lagunas T, Maloney SE, He J, Chen X, Hooda M, Wilkinson MN, 
            Miller TM, Mitra RD, Dougherty JD
  TITLE     A viral toolkit for recording transcription factor-DNA interactions 
            in live mouse tissues.
  JOURNAL   Proc Natl Acad Sci U S A. 2020 May 5;117(18):10003-10014. doi: 
            10.1073/pnas.1918241117. Epub 2020 Apr 16.
  PUBMED    32300008
REFERENCE   2  (bases 1 to 7294)
  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"; doi: 
            "10.1073/pnas.1918241117"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2020-05-5- 5"; volume: "117"; issue: "18"; pages: 
            "10003-10014"
COMMENT     Sequence Label: pAAV_GWRev_Dest
FEATURES             Location/Qualifiers
     source          1..7294
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     22..41
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   183..312
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        413..1591
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          644..1582
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1539..1559
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1622..1655)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(1706..1739)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    1748..1872
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(1897..1902,1903..1920,1921..1927)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             1981..2640
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2047..2066)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             2982..3287
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     3166..3185
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    3461..3494
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    3545..3578
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3603..4191
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3656..3676)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4074..4085)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4223..4699
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4422..4441)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4739..4879
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4739..4868
                     /label=AAV2 ITR
     rep_origin      4954..5409
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5041..5060)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5251..5272
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5426..5445)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5545..5567
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5605..5623)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5691..5795
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5796..5864,5865..6656)
                     /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(6014..6033)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(6827..7294,1..121)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
       61 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
      121 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
      181 gtcctgcagg cagctgcgcg ctcgctcgct cactgaggcc gcccgggcgt cgggcgacct
      241 ttggtcgccc ggcctcagtg agcgagcgag cgcgcagaga gggagtggcc aactccatca
      301 ctaggggttc ctgcggccgc acgcgtaagc tttgcaaaga tggataaagt tttaaacaga
      361 gaggaatctt tgcagctaat ggaccttcta ggtcttgaaa ggagtgggaa ttggctccgg
      421 tgcccgtcag tgggcagagc gcacatcgcc cacagtcccc gagaagttgg ggggaggggt
      481 cggcaattga accggtgcct agagaaggtg gcgcggggta aactgggaaa gtgatgtcgt
      541 gtactggctc cgcctttttc ccgagggtgg gggagaaccg tatataagtg cagtagtcgc
      601 cgtgaacgtt ctttttcgca acgggtttgc cgccagaaca caggtaagtg ccgtgtgtgg
      661 ttcccgcggg cctggcctct ttacgggtta tggcccttgc gtgccttgaa ttacttccac
      721 ctggctgcag tacgtgattc ttgatcccga gcttcgggtt ggaagtgggt gggagagttc
      781 gaggccttgc gcttaaggag ccccttcgcc tcgtgcttga gttgaggcct ggcctgggcg
      841 ctggggccgc cgcgtgcgaa tctggtggca ccttcgcgcc tgtctcgctg ctttcgataa
      901 gtctctagcc atttaaaatt tttgatgacc tgctgcgacg ctttttttct ggcaagatag
      961 tcttgtaaat gcgggccaag atctgcacac tggtatttcg gtttttgggg ccgcgggcgg
     1021 cgacggggcc cgtgcgtccc agcgcacatg ttcggcgagg cggggcctgc gagcgcggcc
     1081 accgagaatc ggacgggggt agtctcaagc tggccggcct gctctggtgc ctggcctcgc
     1141 gccgccgtgt atcgccccgc cctgggcggc aaggctggcc cggtcggcac cagttgcgtg
     1201 agcggaaaga tggccgcttc ccggccctgc tgcagggagc tcaaaatgga ggacgcggcg
     1261 ctcgggagag cgggcgggtg agtcacccac acaaaggaaa agggcctttc cgtcctcagc
     1321 cgtcgcttca tgtgactcca cggagtaccg ggcgccgtcc aggcacctcg attagttctc
     1381 gagcttttgg agtacgtcgt ctttaggttg gggggagggg ttttatgcga tggagtttcc
     1441 ccacactgag tgggtggaga ctgaagttag gccagcttgg cacttgatgt aattctcctt
     1501 ggaatttgcc ctttttgagt ttggatcttg gttcattctc aagcctcaga cagtggttca
     1561 aagttttttt cttccatttc aggtgtcgtg aggtaccgga tcctctagag tcgactccgg
     1621 aataacttcg tataggatac tttatacgaa gttatgcaga atggtagctg gattgtagct
     1681 gctattagca atatgaaacc tcttaataac ttcgtatagc atacattata cgaagttatg
     1741 gcgcgccaca agtttgtaca aaaaagctga acgagaaacg taaaatgata taaatatcaa
     1801 tatattaaat tagattttgc ataaaaaaca gactacataa tactgtaaaa cacaacatat
     1861 ccagtcacta tggcggccgc attaggcacc ccaggcttta cactttatgc ttccggctcg
     1921 tataatgtgt ggattttgag ttaggatccg tcgagatttt caggagctaa ggaagctaaa
     1981 atggagaaaa aaatcactgg atataccacc gttgatatat cccaatggca tcgtaaagaa
     2041 cattttgagg catttcagtc agttgctcaa tgtacctata accagaccgt tcagctggat
     2101 attacggcct ttttaaagac cgtaaagaaa aataagcaca agttttatcc ggcctttatt
     2161 cacattcttg cccgcctgat gaatgctcat ccggaattcc gtatggcaat gaaagacggt
     2221 gagctggtga tatgggatag tgttcaccct tgttacaccg ttttccatga gcaaactgaa
     2281 acgttttcat cgctctggag tgaataccac gacgatttcc ggcagtttct acacatatat
     2341 tcgcaagatg tggcgtgtta cggtgaaaac ctggcctatt tccctaaagg gtttattgag
     2401 aatatgtttt tcgtctcagc caatccctgg gtgagtttca ccagttttga tttaaacgtg
     2461 gccaatatgg acaacttctt cgcccccgtt ttcaccatgg gcaaatatta tacgcaaggc
     2521 gacaaggtgc tgatgccgct ggcgattcag gttcatcatg ccgtttgtga tggcttccat
     2581 gtcggcagaa tgcttaatga attacaacag tactgcgatg agtggcaggg cggggcgtaa
     2641 acgcgtggat ccggcttact aaaagccaga taacagtatg cgtatttgcg cgctgatttt
     2701 tgcggtataa gaatatatac tgatatgtat acccgaagta tgtcaaaaag aggtatgcta
     2761 tgaagcagcg tattacagtg acagttgaca gcgacagcta tcagttgctc aaggcatata
     2821 tgatgtcaat atctccggtc tggtaagcac aaccatgcag aatgaagccc gtcgtctgcg
     2881 tgccgaacgc tggaaagcgg aaaatcagga agggatggct gaggtcgccc ggtttattga
     2941 aatgaacggc tcttttgctg acgagaacag gggctggtga aatgcagttt aaggtttaca
     3001 cctataaaag agagagccgt tatcgtctgt ttgtggatgt acagagtgat attattgaca
     3061 cgcccgggcg acggatggtg atccccctgg ccagtgcacg tctgctgtca gataaagtct
     3121 cccgtgaact ttacccggtg gtgcatatcg gggatgaaag ctggcgcatg atgaccaccg
     3181 atatggccag tgtgccggtc tccgttatcg gggaagaagt ggctgatctc agccaccgcg
     3241 aaaatgacat caaaaacgcc attaacctga tgttctgggg aatataaatg tcaggctccc
     3301 ttatacacag ccagtctgca ggtcgaccat agtgactgga tatgttgtgt tttacagtat
     3361 tatgtagtct gttttttatg caaaatctaa tttaatatat tgatatttat atcattttac
     3421 gtttctcgtt cagctttctt gtacaaattt agttgctagc ataacttcgt ataaagtatc
     3481 ctatacgaag ttatttgcct taacccagaa attatcactg ttattcttta gaatggtgca
     3541 aagaataact tcgtataatg tatgctatac gaagttatga attcgatatc aagcttatcg
     3601 ataatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg
     3661 ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc
     3721 gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt
     3781 tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca
     3841 ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc
     3901 ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc
     3961 tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt ccttggctgc
     4021 tcgcctatgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc
     4081 tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc
     4141 ttcgccttcg ccctcagacg agtcggatct ccctttgggc cgcctccccg catcgatacc
     4201 gagcgctgct cgagagatct acgggtggca tccctgtgac ccctccccag tgcctctcct
     4261 ggccctggaa gttgccactc cagtgcccac cagccttgtc ctaataaaat taagttgcat
     4321 cattttgtct gactaggtgt ccttctataa tattatgggg tggagggggg tggtatggag
     4381 caaggggcaa gttgggaaga caacctgtag ggcctgcggg gtctattggg aaccaagctg
     4441 gagtgcagtg gcacaatctt ggctcactgc aatctccgcc tcctgggttc aagcgattct
     4501 cctgcctcag cctcccgagt tgttgggatt ccaggcatgc atgaccaggc tcagctaatt
     4561 tttgtttttt tggtagagac ggggtttcac catattggcc aggctggtct ccaactccta
     4621 atctcaggtg atctacccac cttggcctcc caaattgctg ggattacagg cgtgaaccac
     4681 tgctcccttc cctgtccttc tgattttgta ggtaaccacg tgcggaccga gcggccgcag
     4741 gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct cactgaggcc
     4801 gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg cggcctcagt gagcgagcga
     4861 gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc tccttacgca tctgtgcggt
     4921 atttcacacc gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg
     4981 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ttagcgcccg
     5041 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     5101 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     5161 aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     5221 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     5281 tcaactctat ctcgggctat tcttttgatt tataagggat tttgccgatt tcggtctatt
     5341 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt
     5401 ttacaatttt atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc
     5461 cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg
     5521 cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat
     5581 caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca
     5641 tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
     5701 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
     5761 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
     5821 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     5881 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
     5941 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
     6001 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     6061 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
     6121 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     6181 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
     6241 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     6301 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
     6361 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
     6421 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     6481 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     6541 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     6601 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
     6661 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     6721 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     6781 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     6841 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     6901 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     6961 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     7021 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     7081 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     7141 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     7201 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     7261 ggtatccggt aagcggcagg gtcggaacag gaga
//
