LOCUS       sequence-437143-        6316 bp    DNA     circular SYN 14-APR-2025
DEFINITION  An adeno-associated viral vector expressing nuclear 
            envelope-embedded eGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6316)
  AUTHORS   Gantz SC, Moussawi K, Hake HS
  TITLE     Delta glutamate receptor conductance drives excitation of mouse 
            dorsal raphe neurons.
  JOURNAL   Elife. 2020 Apr 1;9. pii: 56054. doi: 10.7554/eLife.56054.
  PUBMED    32234214
REFERENCE   2  (bases 1 to 6316)
  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: "Elife."; 
            date: "2020-04-1"; volume: "9. pii: 56054. doi"; pages: " 
            10.7554/eLife.56054"
COMMENT     Sequence Label: pOTTC1553 - pAAV EF1a EGFP-KASH
FEATURES             Location/Qualifiers
     source          1..6316
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        231..1409
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          462..1400
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1357..1377
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      1429..1438
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1435..1437,1438..1440,1441..2151)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1435..1437,1438..1440,1441..2151)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1480..1501)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1741..1760)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2088..2109
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    2443..3031
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2496..2516)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2914..2925)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3063..3539
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3262..3281)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3579..3719
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3579..3708
                     /label=AAV2 ITR
     rep_origin      3794..4249
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3881..3900)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4091..4112
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4266..4285)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4385..4407
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4445..4463)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4531..4635
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4636..4704,4705..5496)
                     /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(4854..4873)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5667..6255
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6156..6175
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtaagctt tgcaaagatg gataaagttt taaacagaga
      181 ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg
      241 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
      301 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
      361 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
      421 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
      481 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct
      541 ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga
      601 ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct
      661 ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt
      721 ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc
      781 ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg
      841 acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac
      901 cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc
      961 cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag
     1021 cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct
     1081 cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg
     1141 tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga
     1201 gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc
     1261 acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg
     1321 aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa
     1381 gtttttttct tccatttcag gtgtcgtgag gtaccggatc caccggtcgc caccatggtg
     1441 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac
     1501 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag
     1561 ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg
     1621 accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac
     1681 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag
     1741 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
     1801 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
     1861 gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc
     1921 aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac
     1981 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     2041 agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     2101 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtccggactc
     2161 agatctcgag aggaggagga ggagacagac agcaggatgc cccacctcga cagccccggc
     2221 agctcccagc cgagacgctc cttcctctca agggtgatca gggcagcgct accgttgcag
     2281 ctgcttctgc tgctgctgct gctcctggcc tgcctgctac ctgcctctga agatgactac
     2341 agctgcaccc aggccaacaa ctttgcccga tccttctacc ccatgctgcg gtacaccaac
     2401 gggccacctc ccacctagga attcgatatc aagcttatcg ataatcaacc tctggattac
     2461 aaaatttgtg aaagattgac tggtattctt aactatgttg ctccttttac gctatgtgga
     2521 tacgctgctt taatgccttt gtatcatgct attgcttccc gtatggcttt cattttctcc
     2581 tccttgtata aatcctggtt gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa
     2641 cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca ctggttgggg cattgccacc
     2701 acctgtcagc tcctttccgg gactttcgct ttccccctcc ctattgccac ggcggaactc
     2761 atcgccgcct gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc
     2821 gtggtgttgt cggggaaatc atcgtccttt ccttggctgc tcgcctatgt tgccacctgg
     2881 attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc ggaccttcct
     2941 tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg
     3001 agtcggatct ccctttgggc cgcctccccg catcgatacc gagcgctgct cgagagatct
     3061 acgggtggca tccctgtgac ccctccccag tgcctctcct ggccctggaa gttgccactc
     3121 cagtgcccac cagccttgtc ctaataaaat taagttgcat cattttgtct gactaggtgt
     3181 ccttctataa tattatgggg tggagggggg tggtatggag caaggggcaa gttgggaaga
     3241 caacctgtag ggcctgcggg gtctattggg aaccaagctg gagtgcagtg gcacaatctt
     3301 ggctcactgc aatctccgcc tcctgggttc aagcgattct cctgcctcag cctcccgagt
     3361 tgttgggatt ccaggcatgc atgaccaggc tcagctaatt tttgtttttt tggtagagac
     3421 ggggtttcac catattggcc aggctggtct ccaactccta atctcaggtg atctacccac
     3481 cttggcctcc caaattgctg ggattacagg cgtgaaccac tgctcccttc cctgtccttc
     3541 tgattttgta ggtaaccacg tgcggaccga gcggccgcag gaacccctag tgatggagtt
     3601 ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg
     3661 acgcccgggc tttgcccggg cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg
     3721 gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgtca
     3781 aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     3841 cgcagcgtga ccgctacact tgccagcgcc ttagcgcccg ctcctttcgc tttcttccct
     3901 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     3961 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgattt gggtgatggt
     4021 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     4081 ttctttaata gtggactctt gttccaaact ggaacaacac tcaactctat ctcgggctat
     4141 tcttttgatt tataagggat tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt
     4201 taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt atggtgcact
     4261 ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc
     4321 gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc
     4381 gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga
     4441 aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag
     4501 acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     4561 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     4621 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     4681 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     4741 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     4801 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     4861 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat
     4921 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     4981 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     5041 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     5101 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     5161 cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa
     5221 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     5281 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     5341 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     5401 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     5461 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     5521 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     5581 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     5641 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     5701 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     5761 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     5821 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     5881 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     5941 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     6001 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     6061 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     6121 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     6181 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     6241 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     6301 ccttttgctc acatgt
//
