LOCUS       sequence-306235-        7999 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Retroviral (MSCV) expression of tdTomato fluorescent protein .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7999)
  AUTHORS   Mann SE, Zhou Z, Landry LG, Anderson AM, Alkanani AK, Fischer J, 
            Peakman M, Mallone R, Campbell K, Michels AW, Nakayama M
  TITLE     Multiplex T Cell Stimulation Assay Utilizing a T Cell Activation 
            Reporter-Based Detection System.
  JOURNAL   Front Immunol. 2020 Apr 9;11:633. doi: 10.3389/fimmu.2020.00633. 
            eCollection 2020.
  PUBMED    32328071
REFERENCE   2  (bases 1 to 7999)
  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: "2020-04-9"; volume: "11:633. doi"; pages: " 
            10.3389/fimmu.2020.00633. eCollection 2020"
COMMENT     Sequence Label: pMIII-Neo-tdTomato
FEATURES             Location/Qualifiers
     source          1..7999
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..517
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
     misc_feature    581..922
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             989..1405
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1335..1357
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1373..1389
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     regulatory      1420..1429
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1426..2856
                     /codon_start=1
                     /product="tandem dimeric (pseudo-monomeric) derivative of 
                     DsRed (Shaner et al., 2004)"
                     /label=tdTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLGHGTGSTGSGSSGTASSEDNNMAVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPY
                     EGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVM
                     NFEDGGLVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGV
                     LKGEIHQALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYE
                     RSEGRHHLFLYGMDELYK"
     misc_feature    2888..3437
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3032..3049)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3259..3278
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3447..4250
                     /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="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3510..3529)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4120..4139
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     LTR             4394..4908
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    5070..5086
                     /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(5094..5100,5101..5118,5119..5124))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5139..5160
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5277..5294)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5448..6036)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5528..5547)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6207..6998,6999..7067))
                     /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     6830..6849
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7068..7172)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7240..7258
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7296..7318)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7418..7437
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7631..7653
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7781..7800
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 aatgaaagac cccacctgta ggtttggcaa gctagcttaa gtaacgccat tttgcaaggc
       61 atggaaaata cataactgag aatagagaag ttcagatcaa ggttaggaac agagagacag
      121 cagaatatgg gccaaacagg atatctgtgg taagcagttc ctgccccggc tcagggccaa
      181 gaacagatgg tccccagatg cggtcccgcc ctcagcagtt tctagagaac catcagatgt
      241 ttccagggtg ccccaaggac ctgaaatgac cctgtgcctt atttgaacta accaatcagt
      301 tcgcttctcg cttctgttcg cgcgcttctg ctccccgagc tcaataaaag agcccacaac
      361 ccctcactcg gcgcgccagt cctccgatag actgcgtcgc ccgggtaccc gtattcccaa
      421 taaagcctct tgctgtttgc atccgaatcg tggactcgct gatccttggg agggtctcct
      481 cagattgatt gactgcccac ctcgggggtc tttcatttgg aggttccacc gagatttgga
      541 gacccctgcc cagggaccac cgaccccccc gccgggaggt aagctggcca gcggtcgttt
      601 cgtgtctgtc tctgtctttg tgcgtgtttg tgccggcatc taatgtttgc gcctgcgtct
      661 gtactagtta gctaactagc tctgtatctg gcggacccgt ggtggaactg acgagttctg
      721 aacacccggc cgcaaccctg ggagacgtcc cagggacttt gggggccgtt tttgtggccc
      781 gacctgagga agggagtcga tgtggaatcc gaccccgtca ggatatgtgg ttctggtagg
      841 agacgagaac ctaaaacagt tcccgcctcc gtctgaattt ttgctttcgg tttggaaccg
      901 aagccgcgcg tcttgtctgc tgcagcgctg cagcatcgtt ctgtgttgtc tctgtctgac
      961 tgtgtttctg tatttgtctg aaaattaggg ccagactgtt accactccct taagtttgac
     1021 cttaggtcac tggaaagatg tcgagcggat cgctcacaac cagtcggtag atgtcaagaa
     1081 gagacgttgg gttaccttct gctctgcaga atggccaacc tttaacgtcg gatggccgcg
     1141 agacggcacc tttaaccgag acctcatcac ccaggttaag atcaaggtct tttcacctgg
     1201 cccgcatgga cacccagacc aggtccccta catcgtgacc tgggaagcct tggcttttga
     1261 cccccctccc tgggtcaagc cctttgtaca ccctaagcct ccgcctcctc ttcctccatc
     1321 cgccccgtct ctcccccttg aacctcctcg ttcgaccccg cctcgatcct ccctttatcc
     1381 agccctcact ccttctctag gcgccggaat tcagatctag ccaccatggt gagcaagggc
     1441 gaggaggtca tcaaagagtt catgcgcttc aaggtgcgca tggagggctc catgaacggc
     1501 cacgagttcg agatcgaggg cgagggcgag ggccgcccct acgagggcac ccagaccgcc
     1561 aagctgaagg tgaccaaggg cggccccctg cccttcgcct gggacatcct gtccccccag
     1621 ttcatgtacg gctccaaggc gtacgtgaag caccccgccg acatccccga ttacaagaag
     1681 ctgtccttcc ccgagggctt caagtgggag cgcgtgatga acttcgagga cggcggtctg
     1741 gtgaccgtga cccaggactc ctccctgcag gacggcacgc tgatctacaa ggtgaagatg
     1801 cgcggcacca acttcccccc cgacggcccc gtaatgcaga agaagaccat gggctgggag
     1861 gcctccaccg agcgcctgta cccccgcgac ggcgtgctga agggcgagat ccaccaggcc
     1921 ctgaagctga aggacggcgg ccactacctg gtggagttca agaccatcta catggccaag
     1981 aagcccgtgc aactgcccgg ctactactac gtggacacca agctggacat cacctcccac
     2041 aacgaggact acaccatcgt ggaacagtac gagcgctccg agggccgcca ccacctgttc
     2101 ctggggcatg gcaccggcag caccggcagc ggcagctccg gcaccgcctc ctccgaggac
     2161 aacaacatgg ccgtcatcaa agagttcatg cgcttcaagg tgcgcatgga gggctccatg
     2221 aacggccacg agttcgagat cgagggcgag ggcgagggcc gcccctacga gggcacccag
     2281 accgccaagc tgaaggtgac caagggcggc cccctgccct tcgcctggga catcctgtcc
     2341 ccccagttca tgtacggctc caaggcgtac gtgaagcacc ccgccgacat ccccgattac
     2401 aagaagctgt ccttccccga gggcttcaag tgggagcgcg tgatgaactt cgaggacggc
     2461 ggtctggtga ccgtgaccca ggactcctcc ctgcaggacg gcacgctgat ctacaaggtg
     2521 aagatgcgcg gcaccaactt cccccccgac ggccccgtaa tgcagaagaa gaccatgggc
     2581 tgggaggcct ccaccgagcg cctgtacccc cgcgacggcg tgctgaaggg cgagatccac
     2641 caggccctga agctgaagga cggcggccac tacctggtgg agttcaagac catctacatg
     2701 gccaagaagc ccgtgcaact gcccggctac tactacgtgg acaccaagct ggacatcacc
     2761 tcccacaacg aggactacac catcgtggaa cagtacgagc gctccgaggg ccgccaccac
     2821 ctgttcctgt acggcatgga cgagctgtac aagtagctcg agcgggatca attccgcccc
     2881 ccccctaacg ttactggccg aagccgcttg gaataaggcc ggtgtgcgtt tgtctatatg
     2941 ttattttcca ccatattgcc gtcttttggc aatgtgaggg cccggaaacc tggccctgtc
     3001 ttcttgacga gcattcctag gggtctttcc cctctcgcca aaggaatgca aggtctgttg
     3061 aatgtcgtga aggaagcagt tcctctggaa gcttcttgaa gacaaacaac gtctgtagcg
     3121 accctttgca ggcagcggaa ccccccacct ggcgacaggt gcctctgcgg ccaaaagcca
     3181 cgtgtataag atacacctgc aaaggcggca caaccccagt gccacgttgt gagttggata
     3241 gttgtggaaa gagtcaaatg gctctcctca agcgtattca acaaggggct gaaggatgcc
     3301 cagaaggtac cccattgtat gggatctgat ctggggcctc ggtgcacatg ctttacatgt
     3361 gtttagtcga ggttaaaaaa acgtctaggc cccccgaacc acggggacgt ggttttcctt
     3421 tgaaaaacac gataatacca tgaaccatgg gatcggccat tgaacaagat ggattgcacg
     3481 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
     3541 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
     3601 tcaagaccga cctgtccggt gccctgaatg aactgcagga cgaggcagcg cggctatcgt
     3661 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
     3721 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
     3781 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
     3841 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
     3901 aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg
     3961 aactgttcgc caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg
     4021 gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact
     4081 gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg
     4141 ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc
     4201 ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gtcgacctgc
     4261 aggcatgcaa gcttcaggta gccggctaac gttaacaacc ggtacctcta gaactatagc
     4321 tagcatgcgc aaatttaaag cgctgatatc gataaaataa aagattttat ttagtctcca
     4381 gaaaaagggg ggaatgaaag accccacctg taggtttggc aagctagctt aagtaacgcc
     4441 attttgcaag gcatggaaaa tacataactg agaatagaga agttcagatc aaggttagga
     4501 acagagagac agcagaatat gggccaaaca ggatatctgt ggtaagcagt tcctgccccg
     4561 gctcagggcc aagaacagat ggtccccaga tgcggtcccg ccctcagcag tttctagaga
     4621 accatcagat gtttccaggg tgccccaagg acctgaaatg accctgtgcc ttatttgaac
     4681 taaccaatca gttcgcttct cgcttctgtt cgcgcgcttc tgctccccga gctcaataaa
     4741 agagcccaca acccctcact cggcgcgcca gtcctccgat agactgcgtc gcccgggtac
     4801 ccgtgtatcc aataaaccct cttgcagttg catccgactt gtggtctcgc tgttccttgg
     4861 gagggtctcc tctgagtgat tgactacccg tcagcggggg tctttcatgg gtaacagttt
     4921 cttgaagttg gagaacaaca ttctgagggt aggagtcgaa tattaagtaa tcctgactca
     4981 attagccact gttttgaatc cacatactcc aatactcctg aaatagttca ttatggacag
     5041 cgcagaaaga gctggggaga attgtgaaat tgttatccgc tcacaattcc acacaacata
     5101 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
     5161 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     5221 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     5281 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     5341 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     5401 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     5461 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     5521 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     5581 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     5641 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     5701 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     5761 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     5821 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     5881 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     5941 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     6001 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     6061 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     6121 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     6181 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     6241 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     6301 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     6361 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     6421 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     6481 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     6541 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     6601 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     6661 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     6721 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     6781 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     6841 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     6901 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     6961 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     7021 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     7081 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     7141 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac
     7201 gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc
     7261 tttcgtctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca gctcccggag
     7321 acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca gggcgcgtca
     7381 gcgggtgttg gcgggtgtcg gggctggctt aactatgcgg catcagagca gattgtactg
     7441 agagtgcacc atatgcggtg tgaaataccg cacagatgcg taaggagaaa ataccgcatc
     7501 aggcgccatt cgccattcag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct
     7561 tcgctattac gccagctggc gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg
     7621 ccagggtttt cccagtcacg acgttgtaaa acgacggcgc aaggaatggt gcatgcaagg
     7681 agatggcgcc caacagtccc ccggccacgg ggcctgccac catacccacg ccgaaacaag
     7741 cgctcatgag cccgaagtgg cgagcccgat cttccccatc ggtgatgtcg gcgatatagg
     7801 cgccagcaac cgcacctgtg gcgccggtga tgccggccac gatgcgtccg gcgtagaggc
     7861 gattagtcca atttgttaaa gacaggatat cagtggtcca ggctctagtt ttgactcaac
     7921 aatatcacca gctgaagcct atagagtacg agccatagat aaaataaaag attttattta
     7981 gtctccagaa aaagggggg
//
