LOCUS       Exported                6402 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  AAV transfer vector with mammalian codon-optimized ChEF-citrine 
            between AAV2 ITR.
ACCESSION   .
VERSION     .
KEYWORDS    AAV-oChEF-citrine
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6402)
  AUTHORS   Lin JY, Lin MZ, Steinbach P, Tsien RY
  TITLE     Characterization of engineered channelrhodopsin variants with 
            improved properties and kinetics.
  JOURNAL   Biophys J. 2009 Mar 4;96(5):1803-14. doi: 10.1016/j.bpj.2008.11.034.
  PUBMED    19254539
REFERENCE   2  (bases 1 to 6402)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6402
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     94..111
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    228..249
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        264..294
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    302..318
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     307..329
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     326..342
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     326..342
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   380..509
                     /label=AAV2 ITR
     promoter        591..1037
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     CDS             2158..2874
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2203..2224)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2464..2483)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2811..2832
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    2911..3499
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2964..2984)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3382..3393)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3502..3518)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3503..3519)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    3548..3669
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3559..3578)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3613..3632
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   3831..3960
                     /label=AAV2 ITR
     primer_bind     complement(3979..3996)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3979..3995)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3988..4010)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4137..4592
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4224..4243)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4434..4455
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4618..4722
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4723..5583
                     /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(4941..4960)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5754..6342
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6243..6262
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
       61 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
      121 gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg
      181 cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtgagttag
      241 ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcgtat gttgtgtgga
      301 attgtgagcg gataacaatt tcacacagga aacagctatg accatgatta cgccagattt
      361 aattaaggcc ttaattaggc tgcgcgctcg ctcgctcact gaggccgccc gggcaaagcc
      421 cgggcgtcgg gcgacctttg gtcgcccggc ctcagtgagc gagcgagcgc gcagagaggg
      481 agtggccaac tccatcacta ggggttcctt gtagttaatg attaacccgc catgctactt
      541 atctacgtag ccatgctcta ggaagatctc tgcagagggc cctgcgtatg agtgcaagtg
      601 ggttttagga ccaggatgag gcggggtggg ggtgcctacc tgacgaccga ccccacccac
      661 tggacaagca cccaaccccc attccccaaa ttgcgcatcc cctatcagag agggggaggg
      721 gaaacaggat gcggcgaggc gcgtgcgcac tgccagcttc agcaccgcgg acagtgcctt
      781 cgcccccgcc tggcggcgcg cgccaccgcc gcctcagcac tgaaggcgcg ctgacgtcac
      841 tcgccggtcc cccgcaaact ccccttcccg gccaccttgg tcgcgtccgc gccgccgccg
      901 gcccagccgg accgcaccac gcgaggcgcg agataggggg gcacgggcgc gaccatctgc
      961 gctgcggcgc cggcgactca gcgctgcctc agtctgcggt gggcagcgga ggagtcgtgt
     1021 cgtgcctgag agcgcagtcg aattcaagct gctagcaagg atccaccggt gaccatggtg
     1081 agcagaagac cctggctgct ggccctggcc ctggccgtgg ccctggccgc cggcagcgcc
     1141 ggcgccagca ccggcagcga cgccaccgtg cccgtggcca cccaggacgg ccccgactac
     1201 gtgttccaca gagcccacga gagaatgctg ttccagacca gctacaccct ggagaacaac
     1261 ggcagcgtga tctgcatccc caacaacggc cagtgcttct gcctggcctg gctgaagagt
     1321 aacggcacca acgccgagaa gctggccgcc aacatcctgc agtggatcac cttcgccctg
     1381 agcgccctgt gcctgatgtt ctacggctac cagacctgga agagtacctg cggctgggag
     1441 gagatctacg tggccaccat cgagatgatc aagttcatca tagagtactt ccacgagttc
     1501 gacgagcccg ccgtgatcta cagcagcaac ggcaacaaga ccgtgtggct gagatacgcc
     1561 gagtggctgc tgacctgccc cgtgatcctg atccacctga gcaacctgac cggcctggcc
     1621 aacgactaca acaagagaac catgggcctg ctggtgagcg acatcggcac catcgtgtgg
     1681 ggcaccaccg ccgccctgag caagggctac gtgagagtga tcttcttcct gatgggcctg
     1741 tgctacggca tctacacctt cttcaacgcc gccaaggtgt acatcgaggc ctaccacacc
     1801 gtgcccaagg gcagatgcag acaggtggtg accggcatgg cctggctgtt cttcgtgagc
     1861 tggggcatgt tccccatcct gttcatcctg ggccccgagg gcttcggcgt gctgagcgtg
     1921 tacggcagca ccgtgggcca caccatcatc gacctgatga gcaagaactg ctggggcctg
     1981 ctgggccact acctgagagt gctgatccac gagcacatcc tgatccacgg cgacatcaga
     2041 aagaccacca agctgaacat cggcggcacc gagatcgagg tggagaccct ggtggaggac
     2101 gaggccgagg ccggcgccgt gaacaagggc accggcaagt acgagagcag cctcgagatg
     2161 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
     2221 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc
     2281 aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
     2341 gtgaccacct tcggctacgg cctgatgtgc ttcgcccgct accccgacca catgaagcag
     2401 cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc
     2461 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
     2521 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
     2581 ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc
     2641 atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac
     2701 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac
     2761 ctgagctacc agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg
     2821 ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagtaggct
     2881 agccccgaat tcgatatcaa gcttatcgat aatcaacctc tggattacaa aatttgtgaa
     2941 agattgactg gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta
     3001 atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa
     3061 tcctggttgc tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg
     3121 tgcactgtgt ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc
     3181 ctttccggga ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc
     3241 cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg
     3301 gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg ccacctggat tctgcgcggg
     3361 acgtccttct gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg
     3421 ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc
     3481 ctttgggccg cctccccgca tcgataccgt cgacctcgac ccgggcggcc gcttcgagca
     3541 gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa
     3601 tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat
     3661 aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggagatgtgg
     3721 gaggtttttt aaagcaagta aaacctctac aaatgtggta aaatcgataa ggatcttcct
     3781 agagcatggc tacgtagata agtagcatgg cgggttaatc attaactaca aggaacccct
     3841 agtgatggag ttggccactc cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc
     3901 aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag
     3961 ccttaattaa cctaattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg
     4021 cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga
     4081 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatgggacgc
     4141 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     4201 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     4261 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     4321 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     4381 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     4441 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     4501 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     4561 gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc
     4621 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     4681 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     4741 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     4801 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     4861 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     4921 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     4981 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     5041 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     5101 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     5161 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     5221 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     5281 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     5341 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     5401 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     5461 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     5521 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     5581 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     5641 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     5701 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     5761 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     5821 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     5881 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     5941 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     6001 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     6061 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     6121 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     6181 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     6241 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     6301 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     6361 tttttacggt tcctggcctt ttgctggcct tttgctcaca tg
//
