LOCUS       sequence-316210-        7083 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses AlACR2 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7083)
  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 7083)
  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: AlACR2_EYFP_pcDNA3.1
FEATURES             Location/Qualifiers
     source          1..7083
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        105..484
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        485..688
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     639..659
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     733..752
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        733..751
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(1705..1707,1708..2421)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1747..1768)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2008..2027)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2355..2376
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2451..2492
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2502..2519
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2542..2559)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2548..2772
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2818..3246
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2905..2924)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3115..3136
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3255..3275)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3260..3589
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3440..3575
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3502..3521
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3656..4450
                     /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(3710..3729)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4320..4339
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4626..4747
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4663..4682)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4717..4736
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4796..4812)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4796..4812)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4809..4831)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4820..4836
                     /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(4844..4850,4851..4868,4869..4874))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4889..4910
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5027..5044)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5198..5786)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5278..5297)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5957..6748,6749..6817))
                     /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     6580..6599
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6818..6922)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6997..7016)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc ttagggttag
       61 gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt gattattgac
      121 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
      181 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      241 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      301 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      361 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      421 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      481 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      541 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      601 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      661 acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca ctgcttactg
      721 gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagt taagcttggt
      781 accgagctcg gatccatgag ctccctggtg ctgggagcag gacagtggtt cagcgagtcc
      841 ctgtctaccc tgaagccacg gctgctgagc gtggacgatg agctgccatc cggcgtgttc
      901 ggagcaaggc tgtgcgtgtt cgtgtgctct gccttcctga tcctgtttta cagcttccag
      961 tttcaggcca agaggtgcaa cctgtccccc atcttcgtgc agctggtgac cgtgacatac
     1021 tattgtatct ctacctttgg cagcacatcc tacctggtgc tgaacgtgga ggaccctgat
     1081 aattctaccc acttccagtt tatgctgggc agatatatct tctggatgct gacatgcccc
     1141 atgatcatca gcaacctgtg cgtgctgctg aacatcctga cccctgacat gctggatctg
     1201 ggccacatca cactgctgat ggtgaaggat atcttcctga tgtgctttgg catcctggga
     1261 gcagcacagt ccaccatggg catcaaggcc gccttcgtga gcgccagctt cctgtttgcc
     1321 gcctggctgg cctacgacat catcaacctg gccgtggcca ggagaaagta cttcatggcc
     1381 caccagtatt gttggaattg gatgatcgtg gtcatcgtgc tgttcatggt gagctggaca
     1441 gtgttccctg tgctgtttgt ggtgggcccc cctctgctgg atatctctag ctccggagca
     1501 gaccagatcg gacacgcagt gggcgatctg ttcgccaaga attgcttcgg ctttgtggtg
     1561 tggtacgtgc ggtttgtggt gctggagcct tataccacac ggaacgccat gcaccgctcc
     1621 gtgtctccat acgccagcaa tctgtctagc aagcagatca agctgggaga gatctccgac
     1681 gaggccctga ggaccgcggc cgccgtgagc aagggcgagg agctgttcac cggggtggtg
     1741 cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag
     1801 ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag
     1861 ctgcccgtgc cctggcccac cctcgtgacc accttcggct acggcctgca gtgcttcgcc
     1921 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac
     1981 gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg
     2041 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag
     2101 gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc
     2161 atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag
     2221 gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc
     2281 gtgctgctgc ccgacaacca ctacctgagc taccagtccg ccctgagcaa agaccccaac
     2341 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc
     2401 atggacgagc tgtacaagta acatatgctc tagagggccc gcggttcgaa ggtaagccta
     2461 tccctaaccc tctcctcggt ctcgattcta cgcgtaccgg tcatcatcac catcaccatt
     2521 gagtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt
     2581 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     2641 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     2701 tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg
     2761 tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg
     2821 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     2881 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     2941 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     3001 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     3061 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     3121 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     3181 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     3241 cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     3301 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     3361 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     3421 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     3481 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct
     3541 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg
     3601 agcttgtata tccattttcg gatctgatca agagacagga tgaggatcgt ttcgcatgat
     3661 tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta
     3721 tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca
     3781 ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga
     3841 cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga
     3901 cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct
     3961 cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg
     4021 gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga
     4081 gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca
     4141 tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga
     4201 ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg
     4261 cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc
     4321 gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt
     4381 gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga
     4441 gttcttctga gcgggactct ggggttcgcg aaatgaccga ccaagcgacg cccaacctgc
     4501 catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     4561 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     4621 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     4681 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     4741 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     4801 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     4861 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     4921 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     4981 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     5041 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     5101 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     5161 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     5221 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     5281 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     5341 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     5401 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     5461 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     5521 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     5581 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     5641 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     5701 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     5761 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     5821 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     5881 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     5941 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     6001 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     6061 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     6121 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     6181 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     6241 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     6301 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     6361 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     6421 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     6481 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     6541 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     6601 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     6661 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     6721 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     6781 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     6841 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     6901 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat
     6961 cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg atgccgcata
     7021 gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa
     7081 aat
//
