LOCUS       sequence-245075-        7233 bp    DNA     circular SYN 14-APR-2025
DEFINITION  sCre-dependent ChR2(H134R)-eYFP with the sLox sites in the reverse 
            orientation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7233)
  TITLE     INTRSECT 2.0
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7233)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pAAV-Ef1a-sCreDIO hChR2(H134R)-eYFP 2.0
FEATURES             Location/Qualifiers
     source          1..7233
                     /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"
     CDS             complement(join(1546..2259,2260..2262,2263..2265))
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1591..1612)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1940..1959
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2199..2220
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      2262..2271
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             complement(2281..3207)
                     /codon_start=1
                     /product="H134R mutant of an N-terminal fragment of 
                     channelrhodopsin-2 from the alga Chlamydomonas reinhardtii 
                     (Nagel et al., 2005)"
                     /label=ChR2(H134R)
                     /note="blue light-activated cation channel"
                     /translation="MDYGGALSAVGRELLFVTNPVVVNGSVLVPEDQCYCAGWIESRGT
                     NGAQTASNVLQWLAAGFSILLLMFYAYQTWKSTCGWEEIYVCAIEMVKVILEFFFEFKN
                     PSMLYLATGHRVQWLRYAEWLLTCPVILIRLSNLTGLSNDYSRRTMGLLVSDIGTIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     regulatory      3204..3213
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     misc_feature    3360..3948
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3413..3433)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3831..3842)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3980..4456
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4179..4198)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4496..4636
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4496..4625
                     /label=AAV2 ITR
     rep_origin      4711..5166
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4798..4817)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5008..5029
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5183..5202)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5302..5324
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5362..5380)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5448..5552
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5553..5621,5622..6413)
                     /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(5771..5790)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6584..7172
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7073..7092
                     /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 gatccactcg tgtccgataa ctgtaattat
     1441 cggacatgat aattgttatt ggtgcaaaga atcacttcct taagtcttta tactgcccag
     1501 ctcgtgtccg ataagtgtat ttatcggaca tgatggcgcg ccctattact tgtacagctc
     1561 gtccatgccg agagtgatcc cggcggcggt cacgaactcc agcaggacca tgtgatcgcg
     1621 cttctcgttg gggtctttgc tcagggcgga ctggtagctc aggtagtggt tgtcgggcag
     1681 cagcacgggg ccgtcgccga tgggggtgtt ctgctggtag tggtcggcga gctgcacgct
     1741 gccgtcctcg atgttgtggc ggatcttgaa gttcaccttg atgccgttct tctgcttgtc
     1801 ggccatgata tagacgttgt ggctgttgta gttgtactcc agcttgtgcc ccaggatgtt
     1861 gccgtcctcc ttgaagtcga tgcccttcag ctcgatgcgg ttcaccaggg tgtcgccctc
     1921 gaacttcacc tcggcgcggg tcttgtagtt gccgtcgtcc ttgaagaaga tggtgcgctc
     1981 ctggacgtag ccttcgggca tggcggactt gaagaagtcg tgctgcttca tgtggtcggg
     2041 gtagcgggcg aagcactgca ggccgtagcc gaaggtggtc acgagggtgg gccagggcac
     2101 gggcagcttg ccggtggtgc agatgaactt cagggtcagc ttgccgtagg tggcatcgcc
     2161 ctcgccctcg ccggacacgc tgaacttgtg gccgtttacg tcgccgtcca gctcgaccag
     2221 gatgggcacc accccggtga acagctcctc gcccttgctc accatggtgg cggccgctgg
     2281 cacggctccg gcctcggctt cgtcttcgac gagagtctcg acctcgatct ccgttccgcc
     2341 gatgttcagt ttggtggttt tgcggatatc tccgtgaatc aatatgtgct cgtggatcag
     2401 gacgcgcagg tagtgtccca acaaccccca acaattttta ctcatcagat caataatcgt
     2461 gtgacctacg gtggagccat agacgctcag gacgccaaaa ccttcgggcc ccaaaatgaa
     2521 gagaattggg aacatacccc agctcacgaa aaacagccat gccatgccgg tcacgacctg
     2581 gcggcaccga ccctttggca cagtatgata accctcgata tatgctttgg cggcgtgaaa
     2641 aaatgtgttc gcgccatagc acaatccaag acaaaagaag atgactttaa catagccggt
     2701 tgccatggcg ctggtagccc cccacacgat agtcccgatg tctgagacaa ggagtcccat
     2761 ggttctcctg ctgtagtcgt tgctcaggcc ggtgaggttg ctcaggcgga taaggatgac
     2821 aggacaagtg agcagccact ctgcatagcg cagccactgc acccggtgtc ctgtggcaag
     2881 gtagagcata gagggattct taaactcaaa aaagaactcg agaatcacct taaccatttc
     2941 aatggcgcac acatagatct cctcccagcc gcatgtagat ttccaggttt ggtaggcata
     3001 gaacatcagc agcaaaatgc tgaatcctgc tgcaagccac tgcaggacat ttgacgcggt
     3061 ctgagcgccg ttcgtgccgc gagattcaat ccatccggca cagtaacatt gatcctcagg
     3121 gaccaggacg gacccgttca ccaccacagg attagtaacg aacaaaagtt cgcgtccgac
     3181 ggcagacaaa gcgccgccat agtccatggt ggctagcatc atgtccgata attacagtta
     3241 tcggacacga gtaagcagac aatcttgtaa tacatgcagc tagtcaagat tgctccagct
     3301 aatcatgtcc gataaataca cttatcggac acgaggaatt cgatatcaag cttatcgata
     3361 atcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac tatgttgctc
     3421 cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt gcttcccgta
     3481 tggctttcat tttctcctcc ttgtataaat cctggttgct gtctctttat gaggagttgt
     3541 ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca acccccactg
     3601 gttggggcat tgccaccacc tgtcagctcc tttccgggac tttcgctttc cccctcccta
     3661 ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt
     3721 tgggcactga caattccgtg gtgttgtcgg ggaaatcatc gtcctttcct tggctgctcg
     3781 cctatgttgc cacctggatt ctgcgcggga cgtccttctg ctacgtccct tcggccctca
     3841 atccagcgga ccttccttcc cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc
     3901 gccttcgccc tcagacgagt cggatctccc tttgggccgc ctccccgcat cgataccgag
     3961 cgctgctcga gagatctacg ggtggcatcc ctgtgacccc tccccagtgc ctctcctggc
     4021 cctggaagtt gccactccag tgcccaccag ccttgtccta ataaaattaa gttgcatcat
     4081 tttgtctgac taggtgtcct tctataatat tatggggtgg aggggggtgg tatggagcaa
     4141 ggggcaagtt gggaagacaa cctgtagggc ctgcggggtc tattgggaac caagctggag
     4201 tgcagtggca caatcttggc tcactgcaat ctccgcctcc tgggttcaag cgattctcct
     4261 gcctcagcct cccgagttgt tgggattcca ggcatgcatg accaggctca gctaattttt
     4321 gtttttttgg tagagacggg gtttcaccat attggccagg ctggtctcca actcctaatc
     4381 tcaggtgatc tacccacctt ggcctcccaa attgctggga ttacaggcgt gaaccactgc
     4441 tcccttccct gtccttctga ttttgtaggt aaccacgtgc ggaccgagcg gccgcaggaa
     4501 cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac tgaggccggg
     4561 cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg
     4621 cgcagctgcc tgcaggggcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt
     4681 tcacaccgca tacgtcaaag caaccatagt acgcgccctg tagcggcgca ttaagcgcgg
     4741 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcctta gcgcccgctc
     4801 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     4861 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     4921 ttgatttggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     4981 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     5041 actctatctc gggctattct tttgatttat aagggatttt gccgatttcg gtctattggt
     5101 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgttta
     5161 caattttatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagcccc
     5221 gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt
     5281 acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac
     5341 cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga
     5401 taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     5461 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     5521 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc
     5581 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     5641 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca
     5701 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt
     5761 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg
     5821 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc
     5881 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     5941 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt
     6001 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     6061 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca
     6121 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg
     6181 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     6241 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     6301 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     6361 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag
     6421 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     6481 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     6541 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     6601 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     6661 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     6721 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     6781 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     6841 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     6901 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     6961 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     7021 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     7081 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     7141 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     7201 tcctggcctt ttgctggcct tttgctcaca tgt
//
