LOCUS       sequence-265100-        6626 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Intersectional viral expression of mCherry in cells expressing Flp 
            AND NOT Cre.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6626)
  AUTHORS   Fenno LE, Ramakrishnan C, Kim YS, Evans KE, Lo M, Vesuna S, Inoue M,
            Cheung KYM, Yuen E, Pichamoorthy N, Hong ASO, Deisseroth K
  TITLE     Comprehensive Dual- and Triple-Feature Intersectional Single-Vector 
            Delivery of Diverse Functional Payloads to Cells of Behaving 
            Mammals.
  JOURNAL   Neuron. 2020 Jun 20. pii: S0896-6273(20)30432-3. doi: 
            10.1016/j.neuron.2020.06.003.
  PUBMED    32574559
REFERENCE   2  (bases 1 to 6626)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Neuron. 
            2020 Jun 20. pii: S0896-6273(20)30432-3. doi: 
            10.1016/j.neuron.2020.06.003."
COMMENT     Sequence Label: pAAV-Ef1a-Coff/Fon-mCherry
FEATURES             Location/Qualifiers
     source          1..6626
                     /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
     promoter        242..1419
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          473..1410
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1367..1387
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1426..1459)
                     /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(1510..1543)
                     /label=loxN
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AGGTATAC) (Shaw et al., 2021). loxN sites are 
                     compatible with each other, but incompatible with loxP or 
                     lox2272 sites."
     regulatory      1550..1559
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1701..1719)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     protein_bind    1918..1951
                     /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    2002..2035
                     /label=loxN
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AGGTATAC) (Shaw et al., 2021). loxN sites are 
                     compatible with each other, but incompatible with loxP or 
                     lox2272 sites."
     primer_bind     complement(2446..2465)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    2753..3341
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2806..2826)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3224..3235)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3373..3849
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3572..3591)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3889..4029
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3889..4018
                     /label=AAV2 ITR
     rep_origin      4104..4559
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4191..4210)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4401..4422
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4576..4595)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4695..4717
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4755..4773)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4841..4945
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4946..5014,5015..5806)
                     /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(5164..5183)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5977..6565
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6466..6485
                     /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 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 tacttccact ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
      601 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
      661 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
      721 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
      781 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
      841 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
      901 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
      961 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
     1021 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
     1081 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
     1141 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
     1201 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     1261 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     1321 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     1381 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gatccataac ttcgtatagg
     1441 atactttata cgaagttatg tagctggagc aatttgaaaa gattgttgta ttacctctag
     1501 ctgcgcttaa taacttcgta tagtatacct tatacgaagt tatactagtg ccaccatggt
     1561 gagcaagggc gaggaggata acatggccat catcaaggag ttcatgcgct tcaaggtgca
     1621 catggagggc tccgtgaacg gccacgagtt cgagatcgag ggcgagggcg agggccgccc
     1681 ctacgagggc acccagaccg ccaagctgaa ggtgaccaag ggtggccccc tgcccttcgc
     1741 ctgggacatc ctgtcccctc agttcatgta cggctccaag gcctacgtga agcaccccgc
     1801 cgacatcccc gactacttga agctgtcctt ccccgagggc ttcaagtggg agcgcgtgat
     1861 gaacttcgag gacggcggcg tggtgaccgt gacccaggtg agtacaggag ggctagcata
     1921 acttcgtata aagtatccta tacgaagtta tttgccttaa cccagaaatt atcactgtta
     1981 ttctttagat aggtgcaaag aataacttcg tataaggtat actatacgaa gttatggtac
     2041 cgaagttcct attcttcaaa tagtatagga acttcaattg ttattggtgc aaagaatcac
     2101 ttccttaagt ctttatactg cccaggaagt tcctattctt caaaaggtat aggaacttcg
     2161 gcgcgcctta cttgtacagc tcgtccatgc cgccggtgga gtggcggccc tcggcgcgtt
     2221 cgtactgttc cacgatggtg tagtcctcgt tgtgggaggt gatgtccaac ttgatgttga
     2281 cgttgtaggc gccgggcagc tgcacgggct tcttggcctt gtaggtggtc ttgacctcag
     2341 cgtcgtagtg gccgccgtcc ttcagcttca gcctctgctt gatctcgccc ttcagggcgc
     2401 cgtcctcggg gtacatccgc tcggaggagg cctcccagcc catggtcttc ttctgcatta
     2461 cggggccgtc ggaggggaag ttggtgccgc gcagcttcac cttgtagatg aactcgccgt
     2521 cctgcaggga ggagtcctgt gggacgacat gacttaacca gttaaccatg ttctctgaac
     2581 atgagaaggg ctggccactc tccagagctc gaagttccta tactatttga agaataggaa
     2641 cttcttgcct taacccagaa attatcactg ttattcttta gaatggtgca aagagaagtt
     2701 cctatacctt ttgaagaata ggaacttcga attcgatatc aagcttatcg ataatcaacc
     2761 tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg ctccttttac
     2821 gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc gtatggcttt
     2881 cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt tgtggcccgt
     2941 tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca ctggttgggg
     3001 cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc ctattgccac
     3061 ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc tgttgggcac
     3121 tgacaattcc gtggtgttgt cggggaaatc atcgtccttt ccttggctgc tcgcctatgt
     3181 tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc
     3241 ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg
     3301 ccctcagacg agtcggatct ccctttgggc cgcctccccg catcgatacc gagcgctgct
     3361 cgagagatct acgggtggca tccctgtgac ccctccccag tgcctctcct ggccctggaa
     3421 gttgccactc cagtgcccac cagccttgtc ctaataaaat taagttgcat cattttgtct
     3481 gactaggtgt ccttctataa tattatgggg tggagggggg tggtatggag caaggggcaa
     3541 gttgggaaga caacctgtag ggcctgcggg gtctattggg aaccaagctg gagtgcagtg
     3601 gcacaatctt ggctcactgc aatctccgcc tcctgggttc aagcgattct cctgcctcag
     3661 cctcccgagt tgttgggatt ccaggcatgc atgaccaggc tcagctaatt tttgtttttt
     3721 tggtagagac ggggtttcac catattggcc aggctggtct ccaactccta atctcaggtg
     3781 atctacccac cttggcctcc caaattgctg ggattacagg cgtgaaccac tgctcccttc
     3841 cctgtccttc tgattttgta ggtaaccacg tgcggaccga gcggccgcag gaacccctag
     3901 tgatggagtt ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa
     3961 aggtcgcccg acgcccgggc tttgcccggg cggcctcagt gagcgagcga gcgcgcagct
     4021 gcctgcaggg gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
     4081 gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     4141 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     4201 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     4261 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgattt
     4321 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     4381 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     4441 ctcgggctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     4501 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt
     4561 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc
     4621 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     4681 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     4741 cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat
     4801 ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     4861 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     4921 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     4981 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     5041 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     5101 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     5161 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
     5221 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     5281 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     5341 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     5401 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     5461 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     5521 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     5581 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     5641 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
     5701 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     5761 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     5821 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     5881 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     5941 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     6001 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     6061 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     6121 tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     6181 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     6241 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     6301 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     6361 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     6421 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     6481 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     6541 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     6601 cttttgctgg ccttttgctc acatgt
//
