LOCUS       Exported                7074 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Highly efficient anion channelrhodopsin from Rhodomonas sp. 
            (CCMP768) for expression in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA3.1-R1ACR_741-EYFP
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7074)
  AUTHORS   Govorunova EG, Sineshchekov OA, Rodarte EM, Janz R, Morelle O, 
            Melkonian M, Wong GK, Spudich JL
  TITLE     The Expanding Family of Natural Anion Channelrhodopsins Reveals 
            Large Variations in Kinetics, Conductance, and Spectral Sensitivity.
  JOURNAL   Sci Rep. 2017 Mar 3;7:43358. doi: 10.1038/srep43358.
  PUBMED    28256618
REFERENCE   2  (bases 1 to 7074)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7074
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(58..78)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        63..392
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      243..378
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     305..324
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             459..1253
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(513..532)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1123..1142
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    1429..1550
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1466..1485)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1520..1539
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1599..1615)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1599..1615)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1612..1634)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1623..1639
                     /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)."
     promoter        complement(1647..1677)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1692..1713
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1830..1847)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2001..2589)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2081..2100)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2760..3620)
                     /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     3383..3402
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3621..3725)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3800..3819)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        3991..4370
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4371..4574
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4525..4545
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4619..4638
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4619..4637
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             5582..6298
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(5624..5645)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5885..5904)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6232..6253
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             6328..6369
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             6379..6396
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6419..6436)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6425..6649
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      6695..49
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6782..6801)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6992..7013
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga
       61 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
      121 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
      181 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
      241 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
      301 tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag
      361 gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt
      421 tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc
      481 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
      541 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
      601 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat
      661 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
      721 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
      781 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
      841 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
      901 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
      961 ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc
     1021 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     1081 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata
     1141 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     1201 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac
     1261 tctggggttc gcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat
     1321 tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg
     1381 atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt
     1441 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     1501 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt
     1561 ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga
     1621 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     1681 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     1741 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     1801 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     1861 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     1921 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     1981 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     2041 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     2101 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     2161 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     2221 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     2281 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     2341 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     2401 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc
     2461 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     2521 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     2581 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     2641 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     2701 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     2761 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     2821 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     2881 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     2941 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     3001 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     3061 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     3121 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     3181 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     3241 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     3301 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     3361 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     3421 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     3481 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     3541 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     3601 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     3661 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     3721 ccgcgcacat ttccccgaaa agtgccacct gacgtcgacg gatcgggaga tctcccgatc
     3781 ccctatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatctgc
     3841 tccctgcttg tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa gctacaacaa
     3901 ggcaaggctt gaccgacaat tgcatgaaga atctgcttag ggttaggcgt tttgcgctgc
     3961 ttcgcgatgt acgggccaga tatacgcgtt gacattgatt attgactagt tattaatagt
     4021 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta
     4081 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga
     4141 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt
     4201 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta
     4261 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg
     4321 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt
     4381 tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc
     4441 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat
     4501 gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct
     4561 atataagcag agctctctgg ctaactagag aacccactgc ttactggctt atcgaaatta
     4621 atacgactca ctatagggag acccaagctg gctagttaag cttggtaccg agctcggatc
     4681 catgtcctac gtggaagatc ctgccttcgg cttttacggc agctgggagg ccgccaatca
     4741 ggactgcatc gagagaggcc tgtccaccag caacaatacc aggatgatcc agctgctgtt
     4801 cgcctccatt tgcgcctgcc aggtgctgtt catggcctcc agggccccta gagtgacctg
     4861 ggaggctgtg tacctgcctg tggtggaaac cgtcctgtac ggcatggcca gcaccggcaa
     4921 cggcaacatg aggctggctg acggcaggat cgtgcctatc gccaggttct gcagctggct
     4981 gatcacctgc cccatcatgc tcttccaagt ggtgggcatc cacgagctga agtggagaac
     5041 catcagctgc aagaacatgg tgatggccgc tagcctgatc aggaccgtgt tcggaatcgg
     5101 cgccagcgtc accatcaccc ccaccatgag gtgggtgcag tacttcctgt ccctgagctt
     5161 cttcctgttc gagctgatcg tggtgtacca gattttctcc gccggcctga agaaattcgg
     5221 cgctgtgaag acctccctga acaacattgt gtactccagg atcatgttcc tgagattcct
     5281 gttcttcctg agctggagca gcttccccat tatctggctc ctgtccagca ccggattctg
     5341 cctggtgaac gaggacgtca gcgtgctggc ctatctggtg gccgacatga tctgcaagaa
     5401 cctgtatggc atcaccaacg ccaacaccct gttcagggtg ctgaatggca agtggacccc
     5461 cgaaaccgag gaagaggcca tgctgcccaa agaccacgaa cacgcccacg tgggcgagac
     5521 ctacttcgag cagatgctga gcggcctcag aggcagcagg aggagggacc acgcggccgc
     5581 cgtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg
     5641 cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg
     5701 caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct
     5761 cgtgaccacc ttcggctacg gcctgcagtg cttcgcccgc taccccgacc acatgaagca
     5821 gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt
     5881 caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt
     5941 gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa
     6001 gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg
     6061 catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga
     6121 ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta
     6181 cctgagctac cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct
     6241 gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagtaaca
     6301 tatgctctag agggcccgcg gttcgaaggt aagcctatcc ctaaccctct cctcggtctc
     6361 gattctacgc gtaccggtca tcatcaccat caccattgag tttaaacccg ctgatcagcc
     6421 tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg
     6481 accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat
     6541 tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag
     6601 gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg
     6661 gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc
     6721 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     6781 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     6841 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     6901 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     6961 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     7021 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcg
//
