LOCUS       Exported                7603 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pDisplay FLIPE-10u
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7603)
  AUTHORS   Okumoto S, Looger LL, Micheva KD, Reimer RJ, Smith SJ, Frommer WB
  TITLE     Detection of glutamate release from neurons by genetically encoded 
            surface-displayed FRET nanosensors.
  JOURNAL   Proc Natl Acad Sci U S A. 2005 Jun 14. 102(24):8740-5.
  PUBMED    15939876
REFERENCE   2  (bases 1 to 7603)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7603
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        10..389
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        390..593
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     544..564
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     638..657
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        638..656
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     sig_peptide     737..799
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
     CDS             800..826
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             854..1570
                     /codon_start=1
                     /product="enhanced CFP"
                     /label=ECFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             2429..3145
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             3152..3181
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             3185..3331
                     /codon_start=1
                     /product="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /label=PDGFR-beta TM domain
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     primer_bind     complement(3353..3370)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3359..3583
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(3715..4303)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3795..3814)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    4632..4679
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     4704..4723
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(4911..5699)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDQEHQGL
                     APAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIALA
                     TRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5022..5041)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5626..5645
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(5734..6094)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5748..5886
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5805..5824)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     6051..6071
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     CDS             complement(6154..7014)
                     /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     6777..6796
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7015..7119)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      7146..7601
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7233..7252)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7443..7464
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 gcgcgcgttg acattgatta ttgactagtt attaatagta atcaattacg gggtcattag
       61 ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct
      121 gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc
      181 caatagggac tttccattga cgtcaatggg tggactattt acggtaaact gcccacttgg
      241 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat
      301 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
      361 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
      421 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
      481 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
      541 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctctctggc
      601 taactagaga acccactgct tactggctta tcgaaattaa tacgactcac tatagggaga
      661 cccaagcttg gtaccgagct cggatccact agtaacggcc gccagtgtgc tggaattcgg
      721 cttggggata tccaccatgg agacagacac actcctgcta tgggtactgc tgctctgggt
      781 tccaggttcc actggtgact atccatatga tgttccagat tatgctgggg cccagccggc
      841 cagatctccc gggatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct
      901 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg
      961 cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt
     1021 gccctggccc accctcgtga ccaccctgac ctggggcgtg cagtgcttca gccgctaccc
     1081 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga
     1141 gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga
     1201 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa
     1261 catcctgggg cacaagctgg agtacaacta catcagccac aacgtctata tcaccgccga
     1321 caagcagaag aacggcatca aggccaactt caagatccgc cacaacatcg aggacggcag
     1381 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     1441 gcccgacaac cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg
     1501 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga
     1561 gctgtacaag ggtggtaccg gaggcgccgc aggcagcacg ctggacaaaa tcgccaaaaa
     1621 cggtgtgatt gtcgtcggtc accgtgaatc ttcagtgcct ttctcttatt acgacaatca
     1681 gcaaaaagtg gtgggttact cgcaggatta ctccaacgcc attgttgaag cagtgaaaaa
     1741 gaaactcaac aaaccggact tgcaggtaaa actgattccg attacctcac aaaaccgtat
     1801 tccactgctg caaaacggca ctttcgattt tgaatgtggt tctaccacca acaacgtcga
     1861 acgccaaaaa caggcggctt tctctgacac tattttcgtg gtcggtacgc gcctgttgac
     1921 caaaaagggt ggcgatatca aagattttgc caacctgaaa gacaaagccg tagtcgtcac
     1981 ttccggcact acctctgaag ttttgctcaa caaactgaat gaagagcaaa aaatgaatat
     2041 gcgcatcatc agcgccaaag atcacggtga ctctttccgc accctggaaa gcggtcgtgc
     2101 cgttgccttt atgatggatg accggctgct ggccggtgaa cgtgcgaaag cgaagaaacc
     2161 agacaactgg gaaatcgtcg gcaagccgca gtctcaggag gcctacggtt gtatgttgcg
     2221 taaagatgat ccgcagttca aaaagctgat ggatgacacc atcgctcagg tgcagacctc
     2281 cggtgaagcg gaaaaatggt ttgataagtg gttcaaaaat ccaattccgc cgaaaaacct
     2341 gaacatgaat ttcgaactgt cagacgaaat gaaagcactg ttcaaagaac cgaatgacaa
     2401 ggcactgaac ggcgccggta ccggtggaat ggtgagcaag ggcgaggagc tgttcaccgg
     2461 ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc
     2521 cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac
     2581 cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ttcggctacg gcctgcagtg
     2641 cttcgcccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga
     2701 aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc
     2761 cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt
     2821 caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca gccacaacgt
     2881 ctatatcatg gccgacaagc agaagaacgg catcaaggtg aacttcaaga tccgccacaa
     2941 catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga
     3001 cggccccgtg ctgctgcccg acaaccacta cctgagctac cagtccgccc tgagcaaaga
     3061 ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac
     3121 tctcggcatg gacgagctgt acaaggtcga cgaacaaaaa ctcatctcag aagaggatct
     3181 gaatgctgtg ggccaggaca cgcaggaggt catcgtggtg ccacactcct tgccctttaa
     3241 ggtggtggtg atctcagcca tcctggccct ggtggtgctc accatcatct cccttatcat
     3301 cctcatcatg ctttggcaga agaagccacg ttaggcggcc gctcgagatc agcctcgact
     3361 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg
     3421 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg
     3481 agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg
     3541 gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga
     3601 accagtggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
     3661 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
     3721 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
     3781 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
     3841 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
     3901 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
     3961 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
     4021 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
     4081 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
     4141 cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
     4201 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
     4261 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
     4321 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
     4381 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
     4441 ctaaagtata tatgagtaac ctgaggctat ggcagggcct gccgccccga cgttggctgc
     4501 gagccctggg ccttcacccg aacttggggg gtggggtggg gaaaaggaag aaacgcgggc
     4561 gtattggccc caatggggtc tcggtggggt atcgacagag tgccagccct gggaccgaac
     4621 cccgcgttta tgaacaaacg acccaacacc gtgcgtttta ttctgtcttt ttattgccgt
     4681 catagcgcgg gttccttccg gtattgtctc cttccgtgtt tcagttagcc tccccctagg
     4741 gtgggcgaag aactccagca tgagatcccc gcgctggagg atcatccagc cggcgtcccg
     4801 gaaaacgatt ccgaagccca acctttcata gaaggcggcg gtggaatcga aatctcgtga
     4861 tggcaggttg ggcgtcgctt ggtcggtcat ttcgaacccc agagtcccgc tcagaagaac
     4921 tcgtcaagaa ggcgatagaa ggcgatgcgc tgcgaatcgg gagcggcgat accgtaaagc
     4981 acgaggaagc ggtcagccca ttcgccgcca agctcttcag caatatcacg ggtagccaac
     5041 gctatgtcct gatagcggtc cgccacaccc agccggccac agtcgatgaa tccagaaaag
     5101 cggccatttt ccaccatgat attcggcaag caggcatcgc catgggtcac gacgagatcc
     5161 tcgccgtcgg gcatgctcgc cttgagcctg gcgaacagtt cggctggcgc gagcccctga
     5221 tgctcttgat catcctgatc gacaagaccg gcttccatcc gagtacgtgc tcgctcgatg
     5281 cgatgtttcg cttggtggtc gaatgggcag gtagccggat caagcgtatg cagccgccgc
     5341 attgcatcag ccatgatgga tactttctcg gcaggagcaa ggtgagatga caggagatcc
     5401 tgccccggca cttcgcccaa tagcagccag tcccttcccg cttcagtgac aacgtcgagc
     5461 acagctgcgc aaggaacgcc cgtcgtggcc agccacgata gccgcgctgc ctcgtcttgc
     5521 agttcattca gggcaccgga caggtcggtc ttgacaaaaa gaaccgggcg cccctgcgct
     5581 gacagccgga acacggcggc atcagagcag ccgattgtct gttgtgccca gtcatagccg
     5641 aatagcctct ccacccaagc ggccggagaa cctgcgtgca atccatcttg ttcaatcatg
     5701 cgaaacgatc ctcatcctgt ctcttgatcg atctttgcaa aagcctaggc ctccaaaaaa
     5761 gcctcctcac tacttctgga atagctcaga ggccgaggag gcggcctcgg cctctgcata
     5821 aataaaaaaa attagtcagc catggggcgg agaatgggcg gaactgggcg gagttagggg
     5881 cgggatgggc ggagttaggg gcgggactat ggttgctgac taattgagat gcatgctttg
     5941 catacttctg cctgctgggg agcctgggga ctttccacac ctggttgctg actaattgag
     6001 atgcatgctt tgcatacttc tgcctgctgg ggagcctggg gactttccac accctaactg
     6061 acacacattc cacagctggt tctttccgcc tcaggactct tcctttttca ataaatcaat
     6121 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
     6181 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
     6241 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
     6301 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
     6361 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
     6421 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
     6481 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
     6541 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
     6601 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
     6661 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
     6721 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
     6781 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
     6841 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
     6901 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
     6961 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
     7021 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
     7081 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
     7141 acctgacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     7201 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct
     7261 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     7321 atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag
     7381 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     7441 tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga
     7501 tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa
     7561 atttaacgcg aattttaaca aaatattaac gcttacaatt tac
//
