LOCUS       Exported                6326 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Expresses an mScarlet-tagged gamma9 subunit for visualizing G 
            beta-gamma translocation.
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA3-mScarlet-gamma9
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6326)
  AUTHORS   Copits BA, O'Neill PR, Gowrishankar R, Yoo JJ, Meshik X, Parker KE, 
            Spangler SM, Vasquez AM, Elerding AJ, Stander MC, Kalyanaraman V, 
            Vogt SK, Samineni VK, Gautam N, Sunahara RK, Gereau RW, Bruchas MR.
  TITLE     A photoswitchable GPCR-based opsin for presynaptic silencing.
  JOURNAL   BioRxiv
REFERENCE   2  (bases 1 to 6326)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6326
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..721
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     672..692
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     766..785
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        766..784
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             820..1515
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet-I
                     /note="matures faster than mScarlet"
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFIKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(953..971)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1221..1240
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             1525..1542
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1820..1837)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1826..2050
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2096..2524
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2183..2202)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2393..2414
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2533..2553)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2538..2867
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2718..2853
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2780..2799
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2934..3728
                     /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(2988..3007)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3598..3617
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3902..4023
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3939..3958)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3993..4012
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4072..4088)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4072..4088)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4085..4107)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4096..4112
                     /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(4120..4150)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4165..4186
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4303..4320)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4474..5062)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4554..4573)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5233..6093)
                     /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     5856..5875
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6094..6198)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6273..6292)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gttggaggtc gctgagtagt gcgcgagcaa aatttaagct acaacaaggc aaggcttgac
       61 cgacaattgc atgaagaatc tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg
      121 ggccagatat acgcgttgac attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacat
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactc
      661 cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      721 tctctggcta actagagaac ccactgctta ctggcttatc gaaattaata cgactcacta
      781 tagggagacc caagctggct agcgtttaaa cttaagctta tggtgagcaa gggcgaggca
      841 gtgatcaagg agttcatgcg gttcaaggtg cacatggagg gctccatgaa cggccacgag
      901 ttcgagatcg agggcgaggg cgagggccgc ccctacgagg gcacccagac cgccaagctg
      961 aaggtgacca agggtggccc cctgcccttc tcctgggaca tcctgtcccc tcagttcatg
     1021 tacggctcca gggccttcat caagcacccc gccgacatcc ccgactacta taagcagtcc
     1081 ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg cgccgtgacc
     1141 gtgacccagg acacctccct ggaggacggc accctgatct acaaggtgaa gctccgcggc
     1201 accaacttcc ctcctgacgg ccccgtaatg cagaagaaga caatgggctg ggaagcgtcc
     1261 accgagcggt tgtaccccga ggacggcgtg ctgaagggcg acattaagat ggccctgcgc
     1321 ctgaaggacg gcggccgcta cctggcggac ttcaagacca cctacaaggc caagaagccc
     1381 gtgcagatgc ccggcgccta caacgtcgac cgcaagttgg acatcacctc ccacaacgag
     1441 gactacaccg tggtggaaca gtacgaacgc tccgagggcc gccactccac cggcggcatg
     1501 gacgagctgt acaagggtac catgcatcac catcaccatc acgcccagga tctcagcgag
     1561 aaggacctgt tgaagatgga ggtggagcag ctgaagaaag aagtgaaaaa cacaagaatt
     1621 ccgatttcca aagcgggaaa ggaaatcaag gagtacgtgg aggcccaagc aggaaacgat
     1681 ccttttctca aaggcatccc tgaggacaag aatcccttca aggaaaaagg cggttgtata
     1741 ataagctgag aattctgcag atatccagca cagtggcggc cgctcgagtc tagagggccc
     1801 gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc
     1861 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa
     1921 aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg
     1981 gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg
     2041 ggctctatgg cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg
     2101 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     2161 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     2221 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     2281 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg
     2341 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     2401 ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg
     2461 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     2521 aattaattct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc tccccagcag
     2581 gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga aagtccccag
     2641 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accatagtcc
     2701 cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc
     2761 atggctgact aatttttttt atttatgcag aggccgaggc cgcctctgcc tctgagctat
     2821 tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctcccgggag
     2881 cttgtatatc cattttcgga tctgatcaag agacaggatg aggatcgttt cgcatgattg
     2941 aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg
     3001 actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg
     3061 ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg
     3121 aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg
     3181 ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc
     3241 tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc
     3301 tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc
     3361 gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc
     3421 aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg
     3481 atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct
     3541 tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt
     3601 tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc
     3661 tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt
     3721 tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc
     3781 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg
     3841 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc
     3901 caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     3961 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     4021 ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct
     4081 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     4141 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     4201 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     4261 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     4321 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     4381 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     4441 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     4501 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     4561 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     4621 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     4681 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     4741 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     4801 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     4861 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     4921 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     4981 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     5041 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     5101 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     5161 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     5221 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     5281 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     5341 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     5401 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     5461 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     5521 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     5581 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     5641 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     5701 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     5761 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     5821 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     5881 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     5941 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     6001 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     6061 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     6121 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     6181 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg acggatcggg
     6241 agatctcccg atcccctatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     6301 agccagtatc tgctccctgc ttgtgt
//
