LOCUS       sequence-100298-        7731 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Human transferrin receptor tagged with pH sensitive orange/red 
            fluorescent protein pHoran4.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7731)
  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 7731)
  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: "10.1083/jcb"; 
            journalName: "J Cell Biol"; date: "2014-11-10- 10"; volume: "207"; 
            issue: "3"; pages: "419-32"
COMMENT     Sequence Label: TfR-pHoran4
FEATURES             Location/Qualifiers
     source          1..7731
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        14..393
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        394..597
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     548..568
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     594..618
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     promoter        828..846
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     828..845
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     859..878
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     CDS             complement(1875..1886)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             3231..3941
                     /codon_start=1
                     /product="orange monomeric derivative of DsRed fluorescent 
                     protein (Shanet et al., 2004)"
                     /label=mOrange
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEENNMAIIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEG
                     FQTAKLKVTKGGPLPFAWDILSPQFTYGSKAYVKHPADIPDYFKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKKRLKLKDGGHYTSEVKTTYKAKKPVQLPGAYIVGIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     3644..3663
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     regulatory      3976..3985
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    3989..4123
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4026..4045)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4080..4099
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     promoter        4196..4553
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4196..4387
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4219..4239)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4404..4539
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(4573..4590)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4573..4589)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4582..4604)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4802..5257
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4889..4908)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5099..5120
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5274..5293)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5428..5450
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5503..5521)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5589..5693
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5694..5762,5763..6554)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMLRTFKVLLCGAVLSRDDAGQEQLGRRIHYSQNDLVDYS
                     PVTEKHLTDGMTVRELCSAAITMSHNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5912..5931)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6725..7313
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7214..7233
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7467..7484
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        join(7637..7642,7643..7660,7661..7667)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7675..7691
                     /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)."
     primer_bind     7680..7702
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7699..7715
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7699..7715
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ttcgagctcg cccgacattg attattgact agttattaat agtaatcaat tacggggtca
       61 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      121 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      181 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      241 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      301 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      361 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
      421 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
      481 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
      541 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagagctcgt
      601 ttagtgaacc gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga
      661 caccgggacc gatccagcct ccgcggccgg gaacggtgca ttggaacgcg gattccccgt
      721 gccaagagtg acgtaagtac cgcctataga gtctataggc ccaccccctt ggcttcgtta
      781 gaacgcggct acaattaata cataacctta tgtatcatac acatacgatt taggtgacac
      841 tatagaataa catccacttt gcctttctct ccacaggtgt ccactcccag gtccaactgc
      901 acctcggttc tatcgattga attccccggg gatccaccat ggatcaagct agatcagcat
      961 tctctaactt gtttggtgga gaaccattgt catatacccg gttcagcctg gctcggcaag
     1021 tagatggcga taacagtcat gtggagatga aacttgctgt agatgaagaa gaaaatgctg
     1081 acaataacac aaaggccaat gtcacaaaac caaaaaggtg tagtggaagt atctgctatg
     1141 ggactattgc tgtgatcgtc tttttcttga ttggatttat gattggctac ttgggctatt
     1201 gtaaaggggt agaaccaaaa actgagtgtg agagactggc aggaaccgag tctccagtga
     1261 gggaggagcc aggagaggac ttccctgcag cacgtcgctt atattgggat gacctgaaga
     1321 gaaagttgtc ggagaaactg gacagcacag acttcaccag caccatcaag ctgctgaatg
     1381 aaaattcata tgtccctcgt gaggctggat ctcaaaaaga tgaaaatctt gcgttgtatg
     1441 ttgaaaatca atttcgtgaa tttaaactca gcaaagtctg gcgtgatcaa cattttgtta
     1501 agattcaggt caaagacagc gctcaaaact cggtgatcat agttgataag aacggtagac
     1561 ttgtttacct ggtggagaat cctgggggtt atgtggcgta tagtaaggct gcaacagtta
     1621 ctggtaaact ggtccatgct aattttggta ctaaaaaaga ttttgaggat ttatacactc
     1681 ctgtgaatgg atctatagtg attgtcagag cagggaaaat cacctttgca gaaaaggttg
     1741 caaatgctga aagcttaaat gcaattggtg tgttgatata catggaccag actaaatttc
     1801 ccattgttaa cgcagaactt tcattctttg gacatgctca tctggggaca ggtgaccctt
     1861 acacacctgg attcccttcc ttcaatcaca ctcagtttcc accatctcgg tcatcaggat
     1921 tgcctaatat acctgtccag acaatctcca gagctgctgc agaaaagctg tttgggaata
     1981 tggaaggaga ctgtccctct gactggaaaa cagactctac atgtaggatg gtaacctcag
     2041 aaagcaagaa tgtgaagctc actgtgagca atgtgctgaa agagataaaa attcttaaca
     2101 tctttggagt tattaaaggc tttgtagaac cagatcacta tgttgtagtt ggggcccaga
     2161 gagatgcatg gggccctgga gctgcaaaat ccggtgtagg cacagctctc ctattgaaac
     2221 ttgcccagat gttctcagat atggtcttaa aagatgggtt tcagcccagc agaagcatta
     2281 tctttgccag ttggagtgct ggagactttg gatcggttgg tgccactgaa tggctagagg
     2341 gatacctttc gtccctgcat ttaaaggctt tcacttatat taatctggat aaagcggttc
     2401 ttggtaccag caacttcaag gtttctgcca gcccactgtt gtatacgctt attgagaaaa
     2461 caatgcaaaa tgtgaagcat ccggttactg ggcaatttct atatcaggac agcaactggg
     2521 ccagcaaagt tgagaaactc actttagaca atgctgcttt ccctttcctt gcatattctg
     2581 gaatcccagc agtttctttc tgtttttgcg aggacacaga ttatccttat ttgggtacca
     2641 ccatggacac ctataaggaa ctgattgaga ggattcctga gttgaacaaa gtggcacgag
     2701 cagctgcaga ggtcgctggt cagttcgtga ttaaactaac ccatgatgtt gaattgaacc
     2761 tggactatga gaggtacaac agccaactgc tttcatttgt gagggatctg aaccaataca
     2821 gagcagacat aaaggaaatg ggcctgagtt tacagtggct gtattctgct cgtggagact
     2881 tcttccgtgc tacttccaga ctaacaacag atttcgggaa tgctgagaaa acagacagat
     2941 ttgtcatgaa gaaactcaat gatcgtgtca tgagagtgga gtatcacttc ctctctccct
     3001 acgtatctcc aaaagagtct cctttccgac atgtcttctg gggctccggc tctcacacgc
     3061 tgccagcttt actggagaac ttgaaactgc gtaaacaaaa taacggtgct tttaatgaaa
     3121 cgctgttcag aaaccagttg gctctagcta cttggactat tcagggagct gcaaatgccc
     3181 tctctggtga cgtttgggac attgacaatg agtttgggga tccaccggtc atggtgagca
     3241 agggcgagga gaataacatg gccatcatca aggagttcat gcgcttcaag gtgcgcatgg
     3301 agggctccgt gaacggccac gagttcgaga tcgagggcga gggcgagggc cgcccctacg
     3361 agggctttca gaccgctaag ctgaaggtga ccaagggtgg ccccctgccc ttcgcctggg
     3421 acatcctgtc ccctcagttc acctacggct ccaaggccta cgtgaagcac cccgccgaca
     3481 tccccgacta cttcaagctg tccttccccg agggcttcaa gtgggagcgc gtgatgaact
     3541 tcgaggacgg cggcgtggtg accgtgaccc aggactcctc cctgcaggac ggcgagttca
     3601 tctacaaggt gaagctgcgc ggcaccaact tcccctccga cggccccgta atgcagaaga
     3661 agaccatggg ctgggaggcc tcctccgagc ggatgtaccc cgaggacggc gccctgaagg
     3721 gcgagatcaa gaagaggctg aagctgaagg acggcggcca ctacacctcc gaggtcaaga
     3781 ccacctacaa ggccaagaag cccgtgcagc tgcccggcgc ctacatcgtc ggcatcaagt
     3841 tggacatcac ctcccacaac gaggactaca ccatcgtgga acagtacgaa cgcgccgagg
     3901 gccgccactc caccggcggc atggacgagc tgtacaagta agctagctaa tctagagtcg
     3961 acctgcagaa gcttggccgc catggcccaa cttgtttatt gcagcttata atggttacaa
     4021 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     4081 tggtttgtcc aaactcatca atgtatctta tcatgtctgg atcgatcggg aattaattcg
     4141 gcgcagcacc atggcctgaa ataacctctg aaagaggaac ttggttaggt accttctgag
     4201 gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc
     4261 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt
     4321 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tgtcaattag tcagcaacca
     4381 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
     4441 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg
     4501 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctgt
     4561 taacagcttg gcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac
     4621 ccaacttaat cgccttgcag cacatccccc cttcgccagc tggcgtaata gcgaagaggc
     4681 ccgcaccgat cgcccttccc aacagttgcg tagcctgaat ggcgaatggc gcctgatgcg
     4741 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atacgtcaaa gcaaccatag
     4801 tacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     4861 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     4921 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     4981 agtgctttac ggcacctcga ccccaaaaaa cttgatttgg gtgatggttc acgtagtggg
     5041 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     5101 ggactcttgt tccaaactgg aacaacactc aaccctatct cgggctattc ttttgattta
     5161 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     5221 aacgcgaatt ttaacaaaat attaacgttt acaattttat ggtgcactct cagtacaatc
     5281 tgctctgatg ccgcatagtt aagccaactc cgctatcgct acgtgactgg gtcatggctg
     5341 cgccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
     5401 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
     5461 catcacccaa acgcgcgagg cagtattctt gaagacgaaa gggcctcgtg atacgcctat
     5521 ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg
     5581 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     5641 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     5701 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     5761 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     5821 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     5881 gttttccaat gctgcgcact tttaaagttc tgctatgtgg cgcggtatta tcccgtgatg
     5941 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgact
     6001 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     6061 ctgccataac catgagtcat aacactgcgg ccaacttact tctgacaacg atcggaggac
     6121 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     6181 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgccagcag
     6241 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     6301 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     6361 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     6421 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     6481 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     6541 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     6601 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     6661 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     6721 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     6781 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     6841 gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc
     6901 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatccgt gtaccagtgg
     6961 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     7021 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     7081 cgacctacac cgaactgaga tacctacagc gtgagcattg agaaagcgcc acgcttcccg
     7141 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     7201 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     7261 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     7321 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     7381 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     7441 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     7501 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatccagc tggcacgaca
     7561 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tacctcactc
     7621 attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga
     7681 gcggataaca atttcacaca ggaaacagct atgaccatga ttacgaatta a
//
