LOCUS       sequence-50205-e        7318 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses NpHR3.0-eYFP in cells lacking Cre (Cre-Off), for 
            optogenetic inhibition.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7318)
  AUTHORS   Saunders A, Johnson CA, Sabatini BL
  TITLE     Novel recombinant adeno-associated viruses for Cre activated and 
            inactivated transgene expression in neurons.
  JOURNAL   Front Neural Circuits. 2012 Jul 27;6:47. doi: 
            10.3389/fncir.2012.00047. eCollection 2012.
  PUBMED    22866029
REFERENCE   2  (bases 1 to 7318)
  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: "Front 
            Neural Circuits."; date: "2012-07-27"; volume: "6:47. doi"; pages: "
            10.3389/fncir.2012.00047. eCollection 2012"
COMMENT     Sequence Label: pAAV-Ef1a-DO-NpHR3.0-eYFP-WPRE-pA
FEATURES             Location/Qualifiers
     source          1..7318
                     /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             join(2537..2539,2540..3250)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2579..2600)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2840..2859)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3187..3208
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    3303..3336
                     /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    3387..3420
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3445..4033
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3498..3518)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3916..3927)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4065..4541
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4264..4283)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4581..4721
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4581..4710
                     /label=AAV2 ITR
     rep_origin      4796..5251
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4883..4902)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5093..5114
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5268..5287)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5387..5409
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5447..5465)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5533..5637
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5638..5706,5707..6498)
                     /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(5856..5875)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6669..7257
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7158..7177
                     /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 gcgccttatc tagatttagc caccatgaca gagaccctgc ctcccgtgac
     1621 cgagagtgcc gtggcccttc aagccgaggt tacccaaagg gagttgttcg agttcgtgct
     1681 gaacgaccct ttgcttgcaa gcagtctcta tatcaacatc gcacttgcag gactgagtat
     1741 actgctgttc gtttttatga cccgaggact cgatgatcca cgggcaaaac ttattgctgt
     1801 gtcaaccatc cttgtgcctg tcgtcagcat tgcctcctac actggattgg cgagcggcct
     1861 gacaatttcc gttcttgaaa tgccagcggg ccattttgca gaaggcagct cagtgatgct
     1921 gggaggagaa gaggtagatg gtgtagtcac catgtgggga cggtatctca cctgggcact
     1981 ttccacgccc atgattctcc tcgctctggg tctcctggcc ggaagcaatg ctacaaagct
     2041 cttcacagct atcactttcg atatcgctat gtgcgtgact ggccttgccg cggccctgac
     2101 tacctcctcc cacctcatga gatggttctg gtacgctatc agttgtgcat gctttctggt
     2161 ggtcttgtat atcctgctgg tggagtgggc acaggacgcc aaagccgcgg gaaccgctga
     2221 catgttcaat accctgaagc tgttgacagt agtgatgtgg ctggggtatc caattgtgtg
     2281 ggctcttgga gtcgagggta tcgcggtgtt gcccgttggg gtgacgagct ggggatattc
     2341 tttcctggat atcgtggcaa agtacatttt cgcattcttg ctcctgaact atctgacgtc
     2401 aaacgaatct gtcgtgtccg gcagcatttt ggatgttcca tctgcttctg ggaccccggc
     2461 tgatgatgcg gccgccaaga gcaggatcac cagcgagggc gagtacatcc ccctggacca
     2521 gatcgacatc aacgtggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct
     2581 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg
     2641 cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt
     2701 gccctggccc accctcgtga ccaccttcgg ctacggcctg cagtgcttcg cccgctaccc
     2761 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga
     2821 gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga
     2881 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa
     2941 catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga
     3001 caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag
     3061 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     3121 gcccgacaac cactacctga gctaccagtc cgccctgagc aaagacccca acgagaagcg
     3181 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga
     3241 gctgtacaag ttctgctacg agaacgaggt gtaaggtggc ggtggcgacc ggtagcgcta
     3301 gcataacttc gtataaagta tcctatacga agttatttgc cttaacccag aaattatcac
     3361 tgttattctt tagaatggtg caaagaataa cttcgtataa tgtatgctat acgaagttat
     3421 gaattcgata tcaagcttat cgataatcaa cctctggatt acaaaatttg tgaaagattg
     3481 actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct
     3541 ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg
     3601 ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact
     3661 gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc
     3721 gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc
     3781 cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa
     3841 tcatcgtcct ttccttggct gctcgcctat gttgccacct ggattctgcg cgggacgtcc
     3901 ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg
     3961 gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg
     4021 gccgcctccc cgcatcgata ccgagcgctg ctcgagagat ctacgggtgg catccctgtg
     4081 acccctcccc agtgcctctc ctggccctgg aagttgccac tccagtgccc accagccttg
     4141 tcctaataaa attaagttgc atcattttgt ctgactaggt gtccttctat aatattatgg
     4201 ggtggagggg ggtggtatgg agcaaggggc aagttgggaa gacaacctgt agggcctgcg
     4261 gggtctattg ggaaccaagc tggagtgcag tggcacaatc ttggctcact gcaatctccg
     4321 cctcctgggt tcaagcgatt ctcctgcctc agcctcccga gttgttggga ttccaggcat
     4381 gcatgaccag gctcagctaa tttttgtttt tttggtagag acggggtttc accatattgg
     4441 ccaggctggt ctccaactcc taatctcagg tgatctaccc accttggcct cccaaattgc
     4501 tgggattaca ggcgtgaacc actgctccct tccctgtcct tctgattttg taggtaacca
     4561 cgtgcggacc gagcggccgc aggaacccct agtgatggag ttggccactc cctctctgcg
     4621 cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg gctttgcccg
     4681 ggcggcctca gtgagcgagc gagcgcgcag ctgcctgcag gggcgcctga tgcggtattt
     4741 tctccttacg catctgtgcg gtatttcaca ccgcatacgt caaagcaacc atagtacgcg
     4801 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     4861 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     4921 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     4981 ttacggcacc tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag tgggccatcg
     5041 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     5101 ttgttccaaa ctggaacaac actcaaccct atctcgggct attcttttga tttataaggg
     5161 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     5221 aattttaaca aaatattaac gtttacaatt ttatggtgca ctctcagtac aatctgctct
     5281 gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc gccctgacgg
     5341 gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg
     5401 tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc
     5461 ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg tggcactttt
     5521 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
     5581 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg
     5641 agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt
     5701 tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga
     5761 gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa
     5821 gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt
     5881 attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt
     5941 gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc
     6001 agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga
     6061 ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat
     6121 cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct
     6181 gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc
     6241 cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg
     6301 gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc
     6361 ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg
     6421 acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca
     6481 ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta gattgattta
     6541 aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc
     6601 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
     6661 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
     6721 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
     6781 actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc
     6841 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
     6901 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
     6961 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
     7021 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
     7081 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
     7141 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
     7201 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
     7261 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgt
//
