LOCUS       sequence-467522-        6996 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expresses NlChR_mCherry in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6996)
  TITLE     Blue-shifted channelrhodopsins from ancyromonads
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6996)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: NlChR_mCherry_pcDNA3.1
FEATURES             Location/Qualifiers
     source          1..6996
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1757..2464
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(1899..1917)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2167..2186
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             2494..2535
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2545..2562
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2585..2602)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2591..2815
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2861..3289
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2948..2967)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3158..3179
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3298..3318)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3303..3632
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3483..3618
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3545..3564
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3699..4493
                     /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(3753..3772)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4363..4382
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4669..4790
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4706..4725)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4760..4779
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4839..4855)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4839..4855)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4852..4874)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4863..4879
                     /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(4887..4893,4894..4911,4912..4917))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4932..4953
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5070..5087)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5241..5829)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5321..5340)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6000..6791,6792..6860))
                     /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     6623..6642
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6861..6965)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagt
      901 taagcttggt accgagctcg gatccgccac catgagctgg gtcgagcata aggccgtgca
      961 gatcagcgcc gagctgctcg tcgtgacagt ctctgtgatt cagctgattg ccaaactgat
     1021 cagaatccgg ggcggaggca gctcctggat cagcgtgtac gtgtgcgcct ctgccgctct
     1081 tcaatacgtg atcagcgtgt cctttctgtt cttcaaccct tttaacgccg gcgccaatgt
     1141 gtacaacgct gaaggcgtgg aagtgtccat ctcaagatac atgttctggt tcctgtcttg
     1201 tcctctgatg gtgctggacc tggtgaagct ggaacacagc gcaaatagac tgtattctca
     1261 gcggctgcca agaaagagcc tggacgagaa gcaggagaaa aaccacggca ccaagatcaa
     1321 ccccgagatc gccctcggcc tggtgctggc catgctgagc tgtgccacaa tgatgagcct
     1381 ggccgctgtt ctgagctgga agctgttctt tctgagcctg ggcggatcta tctgcatggc
     1441 cctgttttac cagttcagcc agatgttcat gctgtacgcc cctgtgctgc ctaacaccat
     1501 ctccaccaaa gccatgatcc ccttcttcta cttcagctgg ctgctgttcc ccaccgtgat
     1561 gtgcctgagc cctatgggct tcaaggtggt gtccttccac gtgagcgagc tgctgttctg
     1621 cctgatcgag ctgcctgcca agaacatcct gctgcagatc atggacaacg tgcagcactc
     1681 tctggataag gaactggctg ctaaggaaca cagcgccagc agcgccctga tcagcaccac
     1741 agccgtggcg gccgccgtga gcaagggcga ggaggataac atggccatca tcaaggagtt
     1801 catgcgcttc aaggtgcaca tggagggctc cgtgaacggc cacgagttcg agatcgaggg
     1861 cgagggcgag ggccgcccct acgagggcac ccagaccgcc aagctgaagg tgaccaaggg
     1921 tggccccctg cccttcgcct gggacatcct gtcccctcag ttcatgtacg gctccaaggc
     1981 ctacgtgaag caccccgccg acatccccga ctacttgaag ctgtccttcc ccgagggctt
     2041 caagtgggag cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga cccaggactc
     2101 ctccctgcag gacggcgagt tcatctacaa ggtgaagctg cgcggcacca acttcccctc
     2161 cgacggcccc gtaatgcaga agaagaccat gggctgggag gcctcctccg agcggatgta
     2221 ccccgaggac ggcgccctga agggcgagat caagcagagg ctgaagctga aggacggcgg
     2281 ccactacgac gctgaggtca agaccaccta caaggccaag aagcccgtgc agctgcccgg
     2341 cgcctacaac gtcaacatca agttggacat cacctcccac aacgaggact acaccatcgt
     2401 ggaacagtac gaacgcgccg agggccgcca ctccaccggc ggcatggacg agctgtacaa
     2461 gtaacatatg ctctagaggg cccgcggttc gaaggtaagc ctatccctaa ccctctcctc
     2521 ggtctcgatt ctacgcgtac cggtcatcat caccatcacc attgagttta aacccgctga
     2581 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct
     2641 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca
     2701 tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag
     2761 ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct
     2821 gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca
     2881 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     2941 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     3001 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     3061 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     3121 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     3181 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     3241 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga
     3301 atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     3361 gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca
     3421 gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc
     3481 ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt
     3541 tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag
     3601 gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt
     3661 tcggatctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc
     3721 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
     3781 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
     3841 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca ggacgaggca gcgcggctat
     3901 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
     3961 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
     4021 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
     4081 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     4141 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     4201 ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg cgaggatctc gtcgtgaccc
     4261 atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct ggattcatcg
     4321 actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct acccgtgata
     4381 ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac ggtatcgccg
     4441 ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc tgagcgggac
     4501 tctggggttc gcgaaatgac cgaccaagcg acgcccaacc tgccatcacg agatttcgat
     4561 tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg
     4621 atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt
     4681 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     4741 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt
     4801 ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga
     4861 aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc
     4921 tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc
     4981 cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc
     5041 ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt
     5101 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca
     5161 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa
     5221 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat
     5281 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc
     5341 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc
     5401 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt
     5461 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac
     5521 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg
     5581 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca
     5641 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc
     5701 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa
     5761 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa
     5821 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac
     5881 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta
     5941 aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt
     6001 taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata
     6061 gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc
     6121 agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac
     6181 cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag
     6241 tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac
     6301 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
     6361 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
     6421 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
     6481 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
     6541 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
     6601 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
     6661 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
     6721 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
     6781 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
     6841 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
     6901 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     6961 ccgcgcacat ttccccgaaa agtgccacct gacgtc
//
