LOCUS       sequence-229248-        7476 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses CD4 fused to TagRFP-T.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7476)
  AUTHORS   Pereira PM, Albrecht D, Culley S, Jacobs C, Marsh M, Mercer J, 
            Henriques R
  TITLE     Fix Your Membrane Receptor Imaging: Actin Cytoskeleton and CD4 
            Membrane Organization Disruption by Chemical Fixation.
  JOURNAL   Front Immunol. 2019 Apr 5;10:675. doi: 10.3389/fimmu.2019.00675. 
            eCollection 2019.
  PUBMED    31024536
REFERENCE   2  (bases 1 to 7476)
  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"; journalName: "Front 
            Immunol."; date: "2019-04-5"; volume: "10:675. doi"; pages: " 
            10.3389/fimmu.2019.00675. eCollection 2019"
COMMENT     Sequence Label: pCD4-TagRFP-T (RHPMP151)
FEATURES             Location/Qualifiers
     source          1..7476
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        116..495
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        496..699
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     650..670
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     744..763
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        744..762
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(808..882,883..2181)
                     /codon_start=1
                     /product="human CD4 cell surface glycoprotein"
                     /label=CD4
                     /translation="MNRGVPFRHLLLVLQLALLPAATQGKKVVLGKKGDTVELTCTASQ
                     KKSIQFHWKNSNQIKILGNQGSFLTKGPSKLNDRADSRRSLWDQGNFPLIIKNLKIEDS
                     DTYICEVEDQKEEVQLLVFGLTANSDTHLLQGQSLTLTLESPPGSSPSVQCRSPRGKNI
                     QGGKTLSVSQLELQDSGTWTCTVLQNQKKVEFKIDIVVLAFQKASSIVYKKEGEQVEFS
                     FPLAFTVEKLTGSGELWWQAERASSSKSWITFDLKNKEVSVKRVTQDPKLQMGKKLPLH
                     LTLPQALPQYAGSGNLTLALEAKTGKLHQEVNLVVMRATQLQKNLTCEVWGPTSPKLML
                     SLKLENKEAKVSKREKAVWVLNPEAGMWQCLLSDSGQVLLESNIKVLPTWSTPVQPMAL
                     IVLGGVAGLLLFIGLGIFFCVRCRHRRRQAERMSQIKRLLSEKKTCQCPHRFQKTCSPI
                     "
     CDS             2197..2910
                     /codon_start=1
                     /product="monomeric derivative of red fluorescent protein 
                     from Entacmaea quadricolor (Merzlyak et al., 2007)"
                     /label=TagRFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTM
                     RIKVVEGGPLPFAFDILATSFMYGSRTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGG
                     VLTATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEANTEMLYPADGGLEGRTD
                     MALKLVGGGHLICNFKTTYRSKKPAKNLKMPGVYYVDHRLERIKEADKETYVEQHEVAV
                     ARYCDLPSKLGHKLN"
     primer_bind     complement(2948..2965)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2954..3178
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3224..3652
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3311..3330)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3521..3542
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3661..3681)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3666..3995
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3846..3981
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3908..3927
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4062..4856
                     /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(4116..4135)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4726..4745
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5030..5151
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5067..5086)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5121..5140
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5200..5216)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5200..5216)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5213..5235)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5224..5240
                     /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(5248..5254,5255..5272,5273..5278))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5293..5314
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5431..5448)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5602..6190)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5682..5701)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6361..7152,7153..7221))
                     /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     6984..7003
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7222..7326)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7401..7420)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat gaagaatctg
       61 cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac gcgttgacat
      121 tgattattga ctagttatta atagtaatca attacggggt cattagttca tagcccatat
      181 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
      241 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
      301 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
      361 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
      421 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
      481 atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt
      541 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
      601 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc
      661 ggtaggcgtg tacggtggga ggtctatata agcagagctc tctggctaac tagagaaccc
      721 actgcttact ggcttatcga aattaatacg actcactata gggagaccca agctggctag
      781 cgtttaaact taagcttagc cgccaccatg aaccggggag tcccttttag gcacttgctt
      841 ctggtgctgc aactggcgct cctcccagca gccactcagg gaaagaaagt ggtgctgggc
      901 aaaaaagggg atacagtgga actgacctgt acagcttccc agaagaagag catacaattc
      961 cactggaaaa actccaacca gataaagatt ctgggaaatc agggctcctt cttaactaaa
     1021 ggtccatcca agctgaatga tcgcgctgac tcaagaagaa gcctttggga ccaaggaaac
     1081 ttccccctga tcatcaagaa tcttaagata gaagactcag atacttacat ctgtgaagtg
     1141 gaggaccaga aggaggaggt gcaattgcta gtgttcggat tgactgccaa ctctgacacc
     1201 cacctgcttc aggggcagag cctgaccctg accttggaga gcccccctgg tagtagcccc
     1261 tcagtgcaat gtaggagtcc aaggggtaaa aacatacagg gggggaagac cctctccgtg
     1321 tctcagctgg agctccagga tagtggcacc tggacatgca ctgtcttgca gaaccagaag
     1381 aaggtggagt tcaaaataga catcgtggtg ctagctttcc agaaggcctc cagcatagtc
     1441 tataagaaag agggggaaca ggtggagttc tccttcccac tcgcctttac agttgaaaag
     1501 ctgacgggca gtggcgagct gtggtggcag gcggagaggg cttcctcctc caagtcttgg
     1561 atcacctttg acctgaagaa caaggaagtg tctgtaaaac gggttaccca ggaccctaag
     1621 ctccagatgg gcaagaagct cccgctccac ctcaccctgc cccaggcctt gcctcagtat
     1681 gctggctctg gaaacctcac cctggccctt gaagcgaaaa caggaaagtt gcatcaggaa
     1741 gtgaacctgg tggtgatgag agccactcag ctccagaaaa atttgacctg tgaggtgtgg
     1801 ggacccacct cccctaagct gatgctgagc ttgaaactgg agaacaagga ggcaaaggtc
     1861 tcgaagcggg agaaggcggt gtgggtgctg aaccctgagg cggggatgtg gcagtgtctg
     1921 ctgagtgact cgggacaggt cctgctggaa tccaacatca aggttctgcc cacatggtcc
     1981 accccggtgc agccaatggc cctgattgtg ctggggggcg tcgccggcct cctgcttttc
     2041 attgggctag gcatcttctt ctgtgtcagg tgccggcacc gaaggcgcca agcagagcgg
     2101 atgtctcaga tcaagagact cctcagtgag aagaagacct gccagtgccc tcaccggttt
     2161 cagaagacat gtagccccat tggggggggg gggtccatgg tgtctaaggg cgaagagctg
     2221 attaaggaga acatgcacat gaagctgtac atggagggca ccgtgaacaa ccaccacttc
     2281 aagtgcacat ccgagggcga aggcaagccc tacgagggca cccagaccat gagaatcaag
     2341 gtggtcgagg gcggccctct ccccttcgcc ttcgacatcc tggctaccag cttcatgtac
     2401 ggcagcagaa ccttcatcaa ccacacccag ggcatccccg acttctttaa gcagtccttc
     2461 cctgagggct tcacatggga gagagtcacc acatacgaag acgggggcgt gctgaccgct
     2521 acccaggaca ccagcctcca ggacggctgc ctcatctaca acgtcaagat cagaggggtg
     2581 aacttcccat ccaacggccc tgtgatgcag aagaaaacac tcggctggga ggccaacacc
     2641 gagatgctgt accccgctga cggcggcctg gaaggcagaa ccgacatggc cctgaagctc
     2701 gtgggcgggg gccacctgat ctgcaacttc aagaccacat acagatccaa gaaacccgct
     2761 aagaacctca agatgcccgg cgtctactat gtggaccaca gactggaaag aatcaaggag
     2821 gccgacaaag agacctacgt cgagcagcac gaggtggctg tggccagata ctgcgacctc
     2881 cctagcaaac tggggcacaa acttaattga ctcgagtcta gagggcccgt ttaaacccgc
     2941 tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg
     3001 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt
     3061 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc
     3121 aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct
     3181 tctgaggcgg aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc
     3241 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     3301 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     3361 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     3421 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     3481 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     3541 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     3601 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt
     3661 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     3721 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     3781 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     3841 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     3901 ttttttttat ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt
     3961 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca
     4021 ttttcggatc tgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat
     4081 tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac
     4141 agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc
     4201 tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaggacgag gcagcgcggc
     4261 tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag
     4321 cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc
     4381 ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg
     4441 atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc
     4501 ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc
     4561 cagccgaact gttcgccagg ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga
     4621 cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca
     4681 tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg
     4741 atattgctga agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg
     4801 ccgctcccga ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgagcgg
     4861 gactctgggg ttcgaaatga ccgaccaagc gacgcccaac ctgccatcac gagatttcga
     4921 ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg
     4981 gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat
     5041 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     5101 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
     5161 tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg
     5221 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
     5281 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
     5341 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     5401 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     5461 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     5521 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     5581 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     5641 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     5701 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     5761 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     5821 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     5881 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     5941 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     6001 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     6061 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     6121 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     6181 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     6241 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     6301 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     6361 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     6421 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     6481 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     6541 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     6601 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     6661 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     6721 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     6781 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     6841 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     6901 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     6961 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     7021 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     7081 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     7141 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     7201 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     7261 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     7321 tccgcgcaca tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atctcccgat
     7381 cccctatggt gcactctcag tacaatctgc tctgatgccg catagttaag ccagtatctg
     7441 ctccctgctt gtgtgttgga ggtcgctgag tagtgc
//
