LOCUS       sequence-316229-        6993 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses strongly red-shifted T1ACR1 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6993)
  AUTHORS   Govorunova EG, Sineshchekov OA, Li H, Wang Y, Brown LS, Spudich JL
  TITLE     RubyACRs, nonalgal anion channelrhodopsins with highly red-shifted 
            absorption.
  JOURNAL   Proc Natl Acad Sci U S A. 2020 Sep 15;117(37):22833-22840. doi: 
            10.1073/pnas.2005981117. Epub 2020 Sep 1.
  PUBMED    32873643
REFERENCE   2  (bases 1 to 6993)
  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"; doi: 
            "10.1073/pnas.2005981117"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2020-09-15- 15"; volume: "117"; issue: "37"; pages: 
            "22833-22840"
COMMENT     Sequence Label: T1ACR1_EYFP_pcDNA3.1
FEATURES             Location/Qualifiers
     source          1..6993
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        98..477
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        478..681
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     632..652
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     726..745
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        726..744
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(1608..1610,1611..2324)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1650..1671)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1911..1930)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2258..2279
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2354..2395
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2405..2422
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2445..2462)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2451..2675
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2721..3149
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2808..2827)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3018..3039
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3158..3178)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3163..3492
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3343..3478
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3405..3424
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3559..4353
                     /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(3613..3632)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4223..4242
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4529..4650
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4566..4585)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4620..4639
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4699..4715)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4699..4715)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4712..4734)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4723..4739
                     /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(4747..4753,4754..4771,4772..4777))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4792..4813
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4930..4947)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5101..5689)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5181..5200)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5860..6651,6652..6720))
                     /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     6483..6502
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6721..6825)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6900..6919)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 acaacaaggc aaggcttgac cgacaattgc atgaagaatc tgcttagggt taggcgtttt
       61 gcgctgcttc gcgatgtacg ggccagatat acgcgttgac attgattatt gactagttat
      121 taatagtaat caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca
      181 taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca
      241 ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg
      301 gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg
      361 ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc
      421 ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg
      481 atgcggtttt ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca
      541 agtctccacc ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt
      601 ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg
      661 gaggtctata taagcagagc tctctggcta actagagaac ccactgctta ctggcttatc
      721 gaaattaata cgactcacta tagggagacc caagctggct agttaagctt ggtaccgagc
      781 tcggatccat ggaccctctg aacatggatg agcacgaggc atggaggaga gacttcgcaa
      841 ggctgctgca ggagcagcct gagagctccg tgccacaggg agtggcagga gccagactgt
      901 gcgtgttttc ttgtagcatc gccctgatct gcttctacgg cgtgcagtat cgggccggca
      961 gatgtaacct gcacccactg tacgtgcagt ttgtgacctt ctgcgtgtat tccctgtctt
     1021 gttttggcac cacagacttc ctgtacatcc caatccagaa tcacgccggc gagacctttg
     1081 atctggccgt gggcaggtat ctgttctgga tgatgacatg ccccgtgatc atctccaacg
     1141 tgaatatcct gctgaacctg atgacccccg acgatctgga gctgcctcgc gtgaccttca
     1201 tgatggtgaa ggacatcgtg atgatcgcct tcggcgtgct ggccgccatg cagaagaacg
     1261 tgaccctgaa gtggatcttt aatctggcct cttacctggt gtgcttctgg ctggtggtgg
     1321 atctgatcca gctgatgaat cagaagaaga agtattttca ggtgcacgag cggtgttggg
     1381 agtggattct ggccacaatg atctacttct ttttcgccca gtctctgttt atcttcctgt
     1441 atgccgtggg ccccccttgt tttaacatca tgggaaggga cggcgatagg atcggacaca
     1501 gcgtgggcga cctgtttgcc aagaatctgt tcggcttttt cgcctggtac gtgaggttcg
     1561 tggtgctgga cccctatgcc aagcggatca gaggagcagc ggccgccgtg agcaagggcg
     1621 aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc
     1681 acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag ctgaccctga
     1741 agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg accaccttcg
     1801 gctacggcct gcagtgcttc gcccgctacc ccgaccacat gaagcagcac gacttcttca
     1861 agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag gacgacggca
     1921 actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac cgcatcgagc
     1981 tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg gagtacaact
     2041 acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc aaggtgaact
     2101 tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac taccagcaga
     2161 acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg agctaccagt
     2221 ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg gagttcgtga
     2281 ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtaacatatg ctctagaggg
     2341 cccgcggttc gaaggtaagc ctatccctaa ccctctcctc ggtctcgatt ctacgcgtac
     2401 cggtcatcat caccatcacc attgagttta aacccgctga tcagcctcga ctgtgccttc
     2461 tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc
     2521 cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg
     2581 tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa
     2641 tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg
     2701 gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     2761 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     2821 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     2881 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     2941 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     3001 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
     3061 ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
     3121 gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca gttagggtgt
     3181 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     3241 gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     3301 ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg
     3361 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
     3421 gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt tggaggccta
     3481 ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga tcaagagaca
     3541 ggatgaggat cgtttcgcat gattgaacaa gatggattgc acgcaggttc tccggccgct
     3601 tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc
     3661 gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc
     3721 ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc cacgacgggc
     3781 gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg
     3841 ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc
     3901 atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac
     3961 caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat
     4021 caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc
     4081 aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg
     4141 aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg
     4201 gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc
     4261 gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc
     4321 gccttctatc gccttcttga cgagttcttc tgagcgggac tctggggttc gcgaaatgac
     4381 cgaccaagcg acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga
     4441 aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga
     4501 tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa
     4561 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     4621 tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta
     4681 gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat
     4741 tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag
     4801 ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg
     4861 ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc
     4921 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     4981 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     5041 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     5101 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     5161 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     5221 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     5281 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     5341 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     5401 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     5461 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     5521 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     5581 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     5641 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     5701 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     5761 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     5821 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     5881 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     5941 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     6001 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     6061 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     6121 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     6181 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     6241 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     6301 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     6361 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     6421 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     6481 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     6541 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     6601 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     6661 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     6721 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     6781 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     6841 agtgccacct gacgtcgacg gatcgggaga tctcccgatc ccctatggtg cactctcagt
     6901 acaatctgct ctgatgccgc atagttaagc cagtatctgc tccctgcttg tgtgttggag
     6961 gtcgctgagt agtgcgcgag caaaatttaa gct
//
