LOCUS       sequence-381584-        6967 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the fusion of monomeric-EGFP-tagged mutant CREB(S133A) in 
            a Cre-dependent manner.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6967)
  AUTHORS   Lee JH, Kim WB, Park EH, Cho JH
  TITLE     Neocortical synaptic engrams for remote contextual memories.
  JOURNAL   Nat Neurosci. 2022 Dec 23. doi: 10.1038/s41593-022-01223-1.
  PUBMED    36564546
REFERENCE   2  (bases 1 to 6967)
  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"; journalName: "Nat 
            Neurosci. 2022 Dec 23. doi: 10.1038/s41593-022-01223-1."
COMMENT     Sequence Label: pAAV-EF1a-DIO-mGFP-MutCREB(S133A)
FEATURES             Location/Qualifiers
     source          1..6967
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     protein_bind    148..168
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     promoter        169..1347
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          400..1338
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1295..1315
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    1347..1371
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(1372..1405)
                     /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(1438..1471)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(join(2528..3238,3239..3241,3242..3244))
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2570..2591)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2919..2938
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3178..3199
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      3241..3250
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3257..3290
                     /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    3323..3356
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(3357..3381)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    3391..3979
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3444..3464)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3862..3873)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4009..4026)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4015..4222
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4230..4370
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4230..4359
                     /label=AAV2 ITR
     rep_origin      4445..4900
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4532..4551)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4742..4763
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4917..4936)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5036..5058
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5096..5114)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5182..5286
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5287..5355,5356..6147)
                     /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(5505..5524)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6318..6906
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6807..6826
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc ttctagacaa ctttgtatag aaaagttggg ctccggtgcc
      181 cgtcagtggg cagagcgcac atcgcccaca gtccccgaga agttgggggg aggggtcggc
      241 aattgaaccg gtgcctagag aaggtggcgc ggggtaaact gggaaagtga tgtcgtgtac
      301 tggctccgcc tttttcccga gggtggggga gaaccgtata taagtgcagt agtcgccgtg
      361 aacgttcttt ttcgcaacgg gtttgccgcc agaacacagg taagtgccgt gtgtggttcc
      421 cgcgggcctg gcctctttac gggttatggc ccttgcgtgc cttgaattac ttccacctgg
      481 ctgcagtacg tgattcttga tcccgagctt cgggttggaa gtgggtggga gagttcgagg
      541 ccttgcgctt aaggagcccc ttcgcctcgt gcttgagttg aggcctggcc tgggcgctgg
      601 ggccgccgcg tgcgaatctg gtggcacctt cgcgcctgtc tcgctgcttt cgataagtct
      661 ctagccattt aaaatttttg atgacctgct gcgacgcttt ttttctggca agatagtctt
      721 gtaaatgcgg gccaagatct gcacactggt atttcggttt ttggggccgc gggcggcgac
      781 ggggcccgtg cgtcccagcg cacatgttcg gcgaggcggg gcctgcgagc gcggccaccg
      841 agaatcggac gggggtagtc tcaagctggc cggcctgctc tggtgcctgg tctcgcgccg
      901 ccgtgtatcg ccccgccctg ggcggcaagg ctggcccggt cggcaccagt tgcgtgagcg
      961 gaaagatggc cgcttcccgg ccctgctgca gggagctcaa aatggaggac gcggcgctcg
     1021 ggagagcggg cgggtgagtc acccacacaa aggaaaaggg cctttccgtc ctcagccgtc
     1081 gcttcatgtg actccacgga gtaccgggcg ccgtccaggc acctcgatta gttctcgagc
     1141 ttttggagta cgtcgtcttt aggttggggg gaggggtttt atgcgatgga gtttccccac
     1201 actgagtggg tggagactga agttaggcca gcttggcact tgatgtaatt ctccttggaa
     1261 tttgcccttt ttgagtttgg atcttggttc attctcaagc ctcagacagt ggttcaaagt
     1321 ttttttcttc catttcaggt gtcgtgacaa gtttgtacaa aaaagcaggc tataacttcg
     1381 tataggatac tttatacgaa gttatcctcg agtaccggtc acgcgtacgc aagcttgata
     1441 acttcgtata gcatacatta tacgaagtta tcctagggcc accttaatct gacttgtggc
     1501 agtaaaggtc cttaagtgct tttagctcct caatcaatgt tttgttttgg ttttcaagca
     1561 ctgccactct gttctctaaa catttcacat attctttctt ctttctacga cattctcttg
     1621 ctgcttccct gttcttcatt agacgaacct ctctctttcg tgctgcttct tcagcaggct
     1681 gtgtaggaag tgctggggag gacgccataa caactccagg ggcaatggtg ctagtgggtg
     1741 ctgtgcgaat ctggtatgtt tgtacatcac cagaggcagc ttgaacaaca acttggttgc
     1801 tgggcactag aatctgctgt ccatcagtgg tctgtgcata ttgtagaatg gtagtacccg
     1861 gctgagtggc agctgcattg gtcatggtta atgtctgcag gccctgtacc ccatcggtac
     1921 cattgttagc cagctgtatt gctcctccct gggtaatggc aatatactgc ccactgctag
     1981 tttggtaaat cggggttggc actgttacag tggtgatggc aggggctgaa gtctcttctt
     2041 ctgatttttc ttcttcaatc cttggcaccc ctggtgcatc agaagataag tcattcaaaa
     2101 ttttcctgta ggcaggcctc cttgaaagga tttcccttcg tttttgggaa tcagttacac
     2161 tatccacaga ctcctgtgaa tcttcacttt ctgcaatagt tgaaatctga gttccggaga
     2221 aaagtctttt taagtcctta caggaagact gaactgtttg gacttgtgga gactgaataa
     2281 ctgatggctg ggccgcctgg atgaccccat ggacctggac tgtctgccca ttgggcagct
     2341 gcactaaggt tacagtggga gcagatgacg tcgcatgagc tgctggcatg gatacctggg
     2401 ctaatgtggc aatctgtggc tgggcttgaa ctgtcatttg ttgactttca gcttctgtta
     2461 cagcagcatc tccactctgc tggttgtctg ctccagagtc catggtcatg gtaggggtag
     2521 gggtaggctt gtacagctcg tccatgccga gagtgatccc ggcggcggtc acgaactcca
     2581 gcaggaccat gtgatcgcgc ttctcgttgg ggtctttgct cagcttggac tgggtgctca
     2641 ggtagtggtt gtcgggcagc agcacggggc cgtcgccgat gggggtgttc tgctggtagt
     2701 ggtcggcgag ctgcacgctg ccgtcctcga tgttgtggcg gatcttgaag ttcaccttga
     2761 tgccgttctt ctgcttgtcg gccatgatat agacgttgtg gctgttgtag ttgtactcca
     2821 gcttgtgccc caggatgttg ccgtcctcct tgaagtcgat gcccttcagc tcgatgcggt
     2881 tcaccagggt gtcgccctcg aacttcacct cggcgcgggt cttgtagttg ccgtcgtcct
     2941 tgaagaagat ggtgcgctcc tggacgtagc cttcgggcat ggcggacttg aagaagtcgt
     3001 gctgcttcat gtggtcgggg tagcggctga agcactgcac gccgtaggtc agggtggtca
     3061 cgagggtggg ccagggcacg ggcagcttgc cggtggtgca gatgaacttc agggtcagct
     3121 tgccgtaggt ggcatcgccc tcgccctcgc cggacacgct gaacttgtgg ccgtttacgt
     3181 cgccgtccag ctcgaccagg atgggcacca ccccggtgaa cagctcctcg cccttgctca
     3241 ccatggtggc tctagaataa cttcgtataa agtatcctat acgaagttat aggtacctga
     3301 gctcaacatc gatagcacta gtataacttc gtataatgta tgctatacga agttataccc
     3361 agctttcttg tacaaagtgg gaattccgat aatcaacctc tggattacaa aatttgtgaa
     3421 agattgactg gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta
     3481 atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa
     3541 tcctggttgc tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg
     3601 tgcactgtgt ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc
     3661 ctttccggga ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc
     3721 cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg
     3781 gggaagctga cgtcctttcc atggctgctc gcctgtgttg ccacctggat tctgcgcggg
     3841 acgtccttct gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg
     3901 ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc
     3961 ctttgggccg cctccccgca tcgggaattc ctagagctcg ctgatcagcc tcgactgtgc
     4021 cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag
     4081 gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta
     4141 ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag
     4201 agaatagcag gcatgctggg gagggccgca ggaaccccta gtgatggagt tggccactcc
     4261 ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc gacgcccggg
     4321 ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg ggcgcctgat
     4381 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgtc aaagcaacca
     4441 tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     4501 accgctacac ttgccagcgc cttagcgccc gctcctttcg ctttcttccc ttcctttctc
     4561 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     4621 tttagtgctt tacggcacct cgaccccaaa aaacttgatt tgggtgatgg ttcacgtagt
     4681 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     4741 agtggactct tgttccaaac tggaacaaca ctcaactcta tctcgggcta ttcttttgat
     4801 ttataaggga ttttgccgat ttcggtctat tggttaaaaa atgagctgat ttaacaaaaa
     4861 tttaacgcga attttaacaa aatattaacg tttacaattt tatggtgcac tctcagtaca
     4921 atctgctctg atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg
     4981 ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg
     5041 agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc
     5101 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt
     5161 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     5221 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     5281 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     5341 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     5401 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     5461 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     5521 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     5581 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     5641 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     5701 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     5761 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     5821 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     5881 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     5941 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     6001 ggaagccgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     6061 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     6121 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     6181 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     6241 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     6301 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     6361 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     6421 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     6481 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     6541 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     6601 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     6661 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     6721 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     6781 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     6841 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     6901 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     6961 cacatgt
//
