LOCUS       sequence-230387-        7104 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid encoding fast anion channelrhodopsin 
            mutant RapACR_T111C_N248Q.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7104)
  TITLE     Mammalian expression plasmid encoding fast anion channelrhodopsin 
            mutant RapACR_T111C_N248Q
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7104)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pcDNA3.1_RapACR_T111C_N248Q-EYFP
FEATURES             Location/Qualifiers
     source          1..7104
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(46..65)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        237..616
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        617..820
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     771..791
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     865..884
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        865..883
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(1858..1860,1861..2574)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1900..1921)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2161..2180)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2508..2529
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2604..2645
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2655..2672
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2695..2712)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2701..2925
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2971..3399
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3058..3077)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3268..3289
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3408..3428)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3413..3742
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3593..3728
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3655..3674
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3809..4603
                     /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(3863..3882)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4473..4492
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4779..4900
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4816..4835)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4870..4889
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4949..4965)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4949..4965)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4962..4984)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4973..4989
                     /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(join(4997..5003,5004..5021,5022..5027))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5042..5063
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5180..5197)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5351..5939)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5431..5450)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6110..6901,6902..6970))
                     /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     6733..6752
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6971..7075)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga
       61 tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg
      121 cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct
      181 gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca
      241 ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc atagcccata
      301 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
      361 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
      421 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
      481 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
      541 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
      601 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
      661 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
      721 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
      781 cggtaggcgt gtacggtggg aggtctatat aagcagagct ctctggctaa ctagagaacc
      841 cactgcttac tggcttatcg aaattaatac gactcactat agggagaccc aagctggcta
      901 gttaagcttg gtaccgagct cggatccatg gactacacca gcctggacga ttttccactg
      961 gagacactgt acttccggaa cggcacagag gtggacccct atgatcacag atgcaagctg
     1021 ggcatccctg gaggctccca gggcatccac acatttcagc tggtgtgggc actgctgtgc
     1081 gcagcacagg ccctgtttct gttctccaag tggccaaaag tgggctggga ggccatgtac
     1141 atgccctgct gtgaggccct gtcttatggc ctgctgagct ccggctacgg ctatatgaag
     1201 ctgcacggcg agttccacta cctgccttgg atcaggtacg ccgcctggct ggtgacctgc
     1261 ccagtgctgc tgaaccaggt gcagggcatg atcgacatca agatcggctc tctgaggctg
     1321 gcaccaatcc agagcctggc aaacctgatc atgatcatca tgggcgtgag cgcccacctg
     1381 caggagaatc acgcctggaa gtggctgttc tttttcgtgg gctgcgtgat gtgcgccgtg
     1441 atctactata tcgcctggct gcagtttgcc aaagtgaatg agctgtacgc cgccaccaag
     1501 acagagatcg gcgataaggt gtgctttcag ctgaagctga tcatgttttg tttctatgtg
     1561 ggctggtcca tgttctctgt gatctggctg ctgtctagcg agggcctgtg cctgatcgag
     1621 gagcacatca gcgccgccgc ccacatccca ggcgacatcc tgagcaagca atcctttggc
     1681 ctgctgatgt ggtataccag gtggatcaca ctgggaggag catgggatat caataagttc
     1741 ctggagaagc aggccaagga gaaggcagag gccgtgctga cccctgccga gaagaggaag
     1801 gccatcatga agcaggccga ggcctgcttc tccatggccg atgcctgtgc ggccgccgtg
     1861 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac
     1921 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag
     1981 ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg
     2041 accaccttcg gctacggcct gcagtgcttc gcccgctacc ccgaccacat gaagcagcac
     2101 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag
     2161 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
     2221 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
     2281 gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc
     2341 aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac
     2401 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     2461 agctaccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     2521 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtaacatatg
     2581 ctctagaggg cccgcggttc gaaggtaagc ctatccctaa ccctctcctc ggtctcgatt
     2641 ctacgcgtac cggtcatcat caccatcacc attgagttta aacccgctga tcagcctcga
     2701 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     2761 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     2821 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     2881 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa
     2941 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg
     3001 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     3061 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     3121 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     3181 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     3241 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     3301 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     3361 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
     3421 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     3481 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     3541 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     3601 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     3661 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
     3721 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
     3781 tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc
     3841 tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg
     3901 ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac
     3961 cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc
     4021 cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg
     4081 gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga
     4141 gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg
     4201 cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg
     4261 tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt
     4321 cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc
     4381 ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg
     4441 gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga
     4501 gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc
     4561 gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc
     4621 gcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg
     4681 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc
     4741 agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata
     4801 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     4861 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga
     4921 cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     4981 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     5041 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     5101 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     5161 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     5221 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     5281 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     5341 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     5401 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     5461 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     5521 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     5581 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     5641 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     5701 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     5761 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     5821 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     5881 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     5941 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     6001 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     6061 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     6121 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     6181 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     6241 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     6301 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     6361 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     6421 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     6481 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     6541 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     6601 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     6661 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     6721 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     6781 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     6841 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     6901 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     6961 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     7021 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     7081 ttccccgaaa agtgccacct gacg
//
