LOCUS       Exported                6015 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Red fluorescent protein based pH indicator in pDisplay vector.
ACCESSION   .
VERSION     .
KEYWORDS    pDisplay-pHuji
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6015)
  AUTHORS   Shen Y, Rosendale M, Campbell RE, Perrais D
  TITLE     pHuji, a pH-sensitive red fluorescent protein for imaging of exo- 
            and endocytosis.
  JOURNAL   J Cell Biol. 2014 Nov 10;207(3):419-32. doi: 10.1083/jcb.201404107.
  PUBMED    25385186
REFERENCE   2  (bases 1 to 6015)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6015
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(86..105)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     296..317
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     enhancer        466..845
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        846..1049
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1000..1020
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1094..1113
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1094..1112
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     sig_peptide     1193..1255
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
     CDS             1256..1282
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1304..2011
                     /codon_start=1
                     /product="pH-sensitive red fluorescent protein derived from
                     mApple, for imaging of exocytosis and endocytosis (Shen et 
                     al., 2014)"
                     /label=pHuji
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEENNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEA
                     FQTAKLKVTKGGPLPFAWDILSPQFMYGSKVYIKHPADIPDYFKLSFPEGFRWERVMNF
                     EDGGIIHVNQDSSLQDGVFIYKVKLRGTNFPSDGPVMQKKTMGWEASEERMYPEDGALK
                     SEIKYRLKLKDGGHYAAEVKTTYKAKKPVQLPGAYIVDIKLDIVSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     1717..1736
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             2021..2050
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2054..2200
                     /codon_start=1
                     /product="transmembrane domain from platelet derived growth
                     factor receptor beta"
                     /label=PDGFR-beta TM domain
                     /translation="AVGQDTQEVIVVPHSLPFKVVVISAILALVVLTIISLIILIMLWQ
                     KKPR"
     primer_bind     complement(2222..2239)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2228..2452
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(2584..3172)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2664..2683)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3501..3548
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     3573..3592
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(3780..4574)
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3891..3910)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4501..4520
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     complement(4673..4692)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     4919..4939
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     CDS             complement(5022..5882)
                     /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     5645..5664
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5883..5987)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6014..454
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
ORIGIN
        1 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
       61 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
      121 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
      181 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca
      241 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
      301 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa
      361 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
      421 gcgaatttta acaaaatatt aacgcttaca atttacgcgc gcgttgacat tgattattga
      481 ctagttatta atagtaatca attacggggt cattagttca tagcccatat atggagttcc
      541 gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat
      601 tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc
      661 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc
      721 caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt
      781 acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta
      841 ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg
      901 gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac
      961 gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg
     1021 tacggtggga ggtctatata agcagagctc tctggctaac tagagaaccc actgcttact
     1081 ggcttatcga aattaatacg actcactata gggagaccca agcttggtac cgagctcgga
     1141 tccactagta acggccgcca gtgtgctgga attcggcttg gggatatcca ccatggagac
     1201 agacacactc ctgctatggg tactgctgct ctgggttcca ggttccactg gtgactatcc
     1261 atatgatgtt ccagattatg ctggggccca gccggccaga tctatggtga gcaagggcga
     1321 ggagaataac atggccatca tcaaggagtt catgcgcttc aaggtgcaca tggagggctc
     1381 cgtgaacggc cacgagttcg agatcgaggg cgagggcgag ggccgcccct acgaggcctt
     1441 tcagaccgct aagctgaagg tgaccaaggg tggccccctg cccttcgcct gggacatcct
     1501 gtcccctcag ttcatgtacg gctccaaggt ctacattaag cacccagccg acatccccga
     1561 ctacttcaag ctgtccttcc ccgagggctt caggtgggag cgcgtgatga acttcgagga
     1621 cggcggcatt attcacgtta accaggactc ctccctgcag gacggcgtgt tcatctacaa
     1681 ggtgaagctg cgcggcacca acttcccctc cgacggcccc gtaatgcaga agaagaccat
     1741 gggctgggag gcctccgagg agcggatgta ccccgaggac ggcgccctga agagcgagat
     1801 taagtatagg ctgaagctga aggacggcgg ccactacgcc gccgaggtca agaccaccta
     1861 caaggccaag aagcccgtgc agctgcccgg cgcctacatc gtggacatca agttggacat
     1921 cgtgtcccac aacgaggact acaccatcgt ggaacagtac gaacgcgccg agggccgcca
     1981 ctccaccggc ggcatggacg agctgtacaa gggcgtcgac gaacaaaaac tcatctcaga
     2041 agaggatctg aatgctgtgg gccaggacac gcaggaggtc atcgtggtgc cacactcctt
     2101 gccctttaag gtggtggtga tctcagccat cctggccctg gtggtgctca ccatcatctc
     2161 ccttatcatc ctcatcatgc tttggcagaa gaagccacgt taggcggccg ctcgagatca
     2221 gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc
     2281 ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg
     2341 cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg
     2401 gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag
     2461 gcggaaagaa ccagtggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     2521 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     2581 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     2641 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     2701 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     2761 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     2821 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     2881 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     2941 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     3001 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     3061 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     3121 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     3181 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     3241 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     3301 taaatcaatc taaagtatat atgagtaacc tgaggctatg gcagggcctg ccgccccgac
     3361 gttggctgcg agccctgggc cttcacccga acttgggggg tggggtgggg aaaaggaaga
     3421 aacgcgggcg tattggcccc aatggggtct cggtggggta tcgacagagt gccagccctg
     3481 ggaccgaacc ccgcgtttat gaacaaacga cccaacaccg tgcgttttat tctgtctttt
     3541 tattgccgtc atagcgcggg ttccttccgg tattgtctcc ttccgtgttt cagttagcct
     3601 ccccctaggg tgggcgaaga actccagcat gagatccccg cgctggagga tcatccagcc
     3661 ggcgtcccgg aaaacgattc cgaagcccaa cctttcatag aaggcggcgg tggaatcgaa
     3721 atctcgtgat ggcaggttgg gcgtcgcttg gtcggtcatt tcgaacccca gagtcccgct
     3781 cagaagaact cgtcaagaag gcgatagaag gcgatgcgct gcgaatcggg agcggcgata
     3841 ccgtaaagca cgaggaagcg gtcagcccat tcgccgccaa gctcttcagc aatatcacgg
     3901 gtagccaacg ctatgtcctg atagcggtcc gccacaccca gccggccaca gtcgatgaat
     3961 ccagaaaagc ggccattttc caccatgata ttcggcaagc aggcatcgcc atgggtcacg
     4021 acgagatcct cgccgtcggg catgctcgcc ttgagcctgg cgaacagttc ggctggcgcg
     4081 agcccctgat gctcttcgtc cagatcatcc tgatcgacaa gaccggcttc catccgagta
     4141 cgtgctcgct cgatgcgatg tttcgcttgg tggtcgaatg ggcaggtagc cggatcaagc
     4201 gtatgcagcc gccgcattgc atcagccatg atggatactt tctcggcagg agcaaggtga
     4261 gatgacagga gatcctgccc cggcacttcg cccaatagca gccagtccct tcccgcttca
     4321 gtgacaacgt cgagcacagc tgcgcaagga acgcccgtcg tggccagcca cgatagccgc
     4381 gctgcctcgt cttgcagttc attcagggca ccggacaggt cggtcttgac aaaaagaacc
     4441 gggcgcccct gcgctgacag ccggaacacg gcggcatcag agcagccgat tgtctgttgt
     4501 gcccagtcat agccgaatag cctctccacc caagcggccg gagaacctgc gtgcaatcca
     4561 tcttgttcaa tcatgcgaaa cgatcctcat cctgtctctt gatcgatctt tgcaaaagcc
     4621 taggcctcca aaaaagcctc ctcactactt ctggaatagc tcagaggccg ggcctcggcc
     4681 tctgcataaa taaaaaaaat tagtcagcca tggggcggag aatgggcgga actgggcgga
     4741 gttaggggcg ggatgggcgg agttaggggc gggactatgg ttgctgacta attgagatgc
     4801 atgctttgca tacttctgcc tgctggggag cctggggact ttccacacct ggttgctgac
     4861 taattgagat gcatgctttg catacttctg cctgctgggg agcctgggga ctttccacac
     4921 cctaactgac acacattcca cagctggttc tttccgcctc aggactcttc ctttttcaat
     4981 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     5041 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     5101 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     5161 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     5221 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     5281 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     5341 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     5401 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     5461 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     5521 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     5581 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     5641 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     5701 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     5761 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     5821 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     5881 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     5941 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     6001 aaagtgccac ctgac
//
