LOCUS       sequence-141982-        6442 bp    DNA     circular SYN 14-APR-2025
DEFINITION  An AAV packaging vector that expresses Cre-dependent 
            nuclear-localized eYFP under control of the EF1a promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6442)
  AUTHORS   Sharpe MJ, Marchant NJ, Whitaker LR, Richie CT, Zhang YJ, Campbell 
            EJ, Koivula PP, Necarsulmer JC, Mejias-Aponte C, Morales M, Pickel 
            J, Smith JC, Niv Y, Shaham Y, Harvey BK, Schoenbaum G
  TITLE     Lateral Hypothalamic GABAergic Neurons Encode Reward Predictions 
            that Are Relayed to the Ventral Tegmental Area to Regulate Learning.
  JOURNAL   Curr Biol. 2017 Jul 24;27(14):2089-2100.e5. doi: 
            10.1016/j.cub.2017.06.024. Epub 2017 Jul 6.
  PUBMED    28690111
REFERENCE   2  (bases 1 to 6442)
  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"; doi: 
            "10.1016/j.cub.2017.06.024"; journalName: "Curr Biol"; date: 
            "2017-07-24- 24"; volume: "27"; issue: "14"; pages: "2089-2100.e5"
COMMENT     Sequence Label: pOTTC1012 - pAAV EF1a DIO Nuc-eYFP
FEATURES             Location/Qualifiers
     source          1..6442
                     /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(1450..1483)
                     /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(1534..1567)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(1588..1608)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(1612..1632)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(1636..1656)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(join(1681..2391,2392..2394,2395..2397))
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1723..1744)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2072..2091
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2331..2352
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     protein_bind    2427..2460
                     /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    2511..2544
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    2569..3157
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2622..2642)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3040..3051)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3189..3665
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3388..3407)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3705..3845
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3705..3834
                     /label=AAV2 ITR
     rep_origin      3920..4375
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4007..4026)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4217..4238
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4392..4411)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4511..4533
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4571..4589)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4657..4761
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4762..4830,4831..5622)
                     /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(4980..4999)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5793..6381
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6282..6301
                     /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 gtaccggatc ctctagagtc
     1441 gactccggaa taacttcgta taggatactt tatacgaagt tatgcagaat ggtagctgga
     1501 ttgtagctgc tattagcaat atgaaacctc ttaataactt cgtatagcat acattatacg
     1561 aagttatggc gcgccgtcga cctatcatac ctttctcttc ttttttggat ctaccttgcg
     1621 cttcttcttg gggtctacct tgcgcttctt ttttgggtcg gcgcggctgc gcaggccgga
     1681 cttgtacagc tcgtccatgc cgagagtgat cccggcggcg gtcacgaact ccagcaggac
     1741 catgtgatcg cgcttctcgt tggggtcttt gctcagggcg gactggtagc tcaggtagtg
     1801 gttgtcgggc agcagcacgg ggccgtcgcc gatgggggtg ttctgctggt agtggtcggc
     1861 gagctgcacg ctgccgtcct cgatgttgtg gcggatcttg aagttcacct tgatgccgtt
     1921 cttctgcttg tcggccatga tatagacgtt gtggctgttg tagttgtact ccagcttgtg
     1981 ccccaggatg ttgccgtcct ccttgaagtc gatgcccttc agctcgatgc ggttcaccag
     2041 ggtgtcgccc tcgaacttca cctcggcgcg ggtcttgtag ttgccgtcgt ccttgaagaa
     2101 gatggtgcgc tcctggacgt agccttcggg catggcggac ttgaagaagt cgtgctgctt
     2161 catgtggtcg gggtagcggg cgaagcactg caggccgtag ccgaaggtgg tcacgagggt
     2221 gggccagggc acgggcagct tgccggtggt gcagatgaac ttcagggtca gcttgccgta
     2281 ggtggcatcg ccctcgccct cgccggacac gctgaacttg tggccgttta cgtcgccgtc
     2341 cagctcgacc aggatgggca ccaccccggt gaacagctcc tcgcccttgc tcaccatggt
     2401 gagatctgag tccggtagcg ctagcgataa cttcgtataa agtatcctat acgaagttat
     2461 ttgccttaac ccagaaatta tcactgttat tctttagaat ggtgcaaaga ataacttcgt
     2521 ataatgtatg ctatacgaag ttatgaattc gatatcaagc ttatcgataa tcaacctctg
     2581 gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta
     2641 tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt
     2701 ttctcctcct tgtataaatc ctggttgctg tctctttatg aggagttgtg gcccgttgtc
     2761 aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg ttggggcatt
     2821 gccaccacct gtcagctcct ttccgggact ttcgctttcc ccctccctat tgccacggcg
     2881 gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac
     2941 aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc ctatgttgcc
     3001 acctggattc tgcgcgggac gtccttctgc tacgtccctt cggccctcaa tccagcggac
     3061 cttccttccc gcggcctgct gccggctctg cggcctcttc cgcgtcttcg ccttcgccct
     3121 cagacgagtc ggatctccct ttgggccgcc tccccgcatc gataccgagc gctgctcgag
     3181 agatctacgg gtggcatccc tgtgacccct ccccagtgcc tctcctggcc ctggaagttg
     3241 ccactccagt gcccaccagc cttgtcctaa taaaattaag ttgcatcatt ttgtctgact
     3301 aggtgtcctt ctataatatt atggggtgga ggggggtggt atggagcaag gggcaagttg
     3361 ggaagacaac ctgtagggcc tgcggggtct attgggaacc aagctggagt gcagtggcac
     3421 aatcttggct cactgcaatc tccgcctcct gggttcaagc gattctcctg cctcagcctc
     3481 ccgagttgtt gggattccag gcatgcatga ccaggctcag ctaatttttg tttttttggt
     3541 agagacgggg tttcaccata ttggccaggc tggtctccaa ctcctaatct caggtgatct
     3601 acccaccttg gcctcccaaa ttgctgggat tacaggcgtg aaccactgct cccttccctg
     3661 tccttctgat tttgtaggta accacgtgcg gaccgagcgg ccgcaggaac ccctagtgat
     3721 ggagttggcc actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt
     3781 cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc gcagctgcct
     3841 gcaggggcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat
     3901 acgtcaaagc aaccatagta cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     3961 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     4021 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     4081 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgatttgggt
     4141 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     4201 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     4261 ggctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     4321 ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgtttac aattttatgg
     4381 tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagccccg acacccgcca
     4441 acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct
     4501 gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg
     4561 agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt
     4621 tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt
     4681 ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa
     4741 taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt
     4801 tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat
     4861 gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag
     4921 atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg
     4981 ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata
     5041 cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat
     5101 ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc
     5161 aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg
     5221 ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac
     5281 gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact
     5341 ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa
     5401 gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct
     5461 ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc
     5521 tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga
     5581 cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac
     5641 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
     5701 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     5761 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     5821 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     5881 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc
     5941 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     6001 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     6061 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     6121 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     6181 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     6241 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     6301 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     6361 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     6421 tgctggcctt ttgctcacat gt
//
