LOCUS       sequence-306229-        7282 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Retroviral (MSCV) expression of ametrine fluorescent protein .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7282)
  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 7282)
  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-AM
FEATURES             Location/Qualifiers
     source          1..7282
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    189..530
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     CDS             597..1013
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     943..965
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     981..997
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     regulatory      1022..1031
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1028..1030,1031..1033,1034..1747)
                     /codon_start=1
                     /product="photostable yellow fluorescent protein with a 
                     large Stokes shift, excitable with violet light (Ai et al.,
                     2008)"
                     /label=mAmetrine1.1
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHRFSVRGEGEGDATNGKLTL
                     KFICTSGELPVPWPTLVTTLSYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDD
                     GSYRTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNMNAWDAYITADKQKNGIK
                     ANFKIEHNVKDGGVQLADAYQQNTPIGDGSVLLPDNHYLSFQSKLFKDPNEQRDHMVLL
                     EFVTAAGITPGMDELYK"
     primer_bind     complement(1073..1094)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     misc_feature    1779..2328
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(1923..1940)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2150..2169
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             2338..3141
                     /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(2401..2420)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3011..3030
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     LTR             3285..3799
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    3961..3977
                     /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(3985..3991,3992..4009,4010..4015))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4030..4051
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4168..4185)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4339..4927)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4419..4438)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5098..5889,5890..5958))
                     /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     5721..5740
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5959..6063)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6131..6149
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6187..6209)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6309..6328
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6522..6544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6672..6691
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     LTR             join(6891..7282,1..125)
                     /label=5' LTR
                     /note="5' long terminal repeat from murine embryonic stem 
                     cell virus"
ORIGIN
        1 tgcgtcgccc gggtacccgt attcccaata aagcctcttg ctgtttgcat ccgaatcgtg
       61 gactcgctga tccttgggag ggtctcctca gattgattga ctgcccacct cgggggtctt
      121 tcatttggag gttccaccga gatttggaga cccctgccca gggaccaccg acccccccgc
      181 cgggaggtaa gctggccagc ggtcgtttcg tgtctgtctc tgtctttgtg cgtgtttgtg
      241 ccggcatcta atgtttgcgc ctgcgtctgt actagttagc taactagctc tgtatctggc
      301 ggacccgtgg tggaactgac gagttctgaa cacccggccg caaccctggg agacgtccca
      361 gggactttgg gggccgtttt tgtggcccga cctgaggaag ggagtcgatg tggaatccga
      421 ccccgtcagg atatgtggtt ctggtaggag acgagaacct aaaacagttc ccgcctccgt
      481 ctgaattttt gctttcggtt tggaaccgaa gccgcgcgtc ttgtctgctg cagcgctgca
      541 gcatcgttct gtgttgtctc tgtctgactg tgtttctgta tttgtctgaa aattagggcc
      601 agactgttac cactccctta agtttgacct taggtcactg gaaagatgtc gagcggatcg
      661 ctcacaacca gtcggtagat gtcaagaaga gacgttgggt taccttctgc tctgcagaat
      721 ggccaacctt taacgtcgga tggccgcgag acggcacctt taaccgagac ctcatcaccc
      781 aggttaagat caaggtcttt tcacctggcc cgcatggaca cccagaccag gtcccctaca
      841 tcgtgacctg ggaagccttg gcttttgacc cccctccctg ggtcaagccc tttgtacacc
      901 ctaagcctcc gcctcctctt cctccatccg ccccgtctct cccccttgaa cctcctcgtt
      961 cgaccccgcc tcgatcctcc ctttatccag ccctcactcc ttctctaggc gccggaattc
     1021 agccaccatg gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga
     1081 gctggacggc gacgtaaacg gccacaggtt cagcgtgcgc ggcgagggcg agggcgatgc
     1141 caccaacggc aagctgaccc tgaagttcat ctgcacctcc ggcgagctgc ccgtgccctg
     1201 gcccaccctc gtgaccaccc tgtcttacgg cgtgcagtgc ttcgcccgct accccgacca
     1261 catgaagcag cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac
     1321 catctccttc aaggacgacg gcagctacag gacccgcgcc gaggtgaagt tcgagggcga
     1381 caccctggtg aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct
     1441 ggggcacaag ctggagtaca acatgaacgc gtgggacgcg tatatcacgg ccgacaagca
     1501 gaagaacggc atcaaagcga acttcaagat cgagcacaac gtcaaggacg gcggcgtgca
     1561 gctcgccgac gcgtaccagc agaacacccc catcggcgac ggctccgtgc tgctgcctga
     1621 caaccactac ctgagcttcc agagcaagct gttcaaagac cccaacgagc agcgcgatca
     1681 catggtcctg ctggagttcg ttaccgccgc cgggataact cccggcatgg acgagctgta
     1741 caagtaactc gagcgggatc aattccgccc cccccctaac gttactggcc gaagccgctt
     1801 ggaataaggc cggtgtgcgt ttgtctatat gttattttcc accatattgc cgtcttttgg
     1861 caatgtgagg gcccggaaac ctggccctgt cttcttgacg agcattccta ggggtctttc
     1921 ccctctcgcc aaaggaatgc aaggtctgtt gaatgtcgtg aaggaagcag ttcctctgga
     1981 agcttcttga agacaaacaa cgtctgtagc gaccctttgc aggcagcgga accccccacc
     2041 tggcgacagg tgcctctgcg gccaaaagcc acgtgtataa gatacacctg caaaggcggc
     2101 acaaccccag tgccacgttg tgagttggat agttgtggaa agagtcaaat ggctctcctc
     2161 aagcgtattc aacaaggggc tgaaggatgc ccagaaggta ccccattgta tgggatctga
     2221 tctggggcct cggtgcacat gctttacatg tgtttagtcg aggttaaaaa aacgtctagg
     2281 ccccccgaac cacggggacg tggttttcct ttgaaaaaca cgataatacc atgaaccatg
     2341 ggatcggcca ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg
     2401 ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg
     2461 ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat
     2521 gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca
     2581 gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg
     2641 gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat
     2701 gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa
     2761 catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg
     2821 gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg
     2881 cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg
     2941 gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat
     3001 caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac
     3061 cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc
     3121 cttcttgacg agttcttctg agtcgacctg caggcatgca agcttcaggt agccggctaa
     3181 cgttaacaac cggtacctct agaactatag ctagcatgcg caaatttaaa gcgctgatat
     3241 cgataaaata aaagatttta tttagtctcc agaaaaaggg gggaatgaaa gaccccacct
     3301 gtaggtttgg caagctagct taagtaacgc cattttgcaa ggcatggaaa atacataact
     3361 gagaatagag aagttcagat caaggttagg aacagagaga cagcagaata tgggccaaac
     3421 aggatatctg tggtaagcag ttcctgcccc ggctcagggc caagaacaga tggtccccag
     3481 atgcggtccc gccctcagca gtttctagag aaccatcaga tgtttccagg gtgccccaag
     3541 gacctgaaat gaccctgtgc cttatttgaa ctaaccaatc agttcgcttc tcgcttctgt
     3601 tcgcgcgctt ctgctccccg agctcaataa aagagcccac aacccctcac tcggcgcgcc
     3661 agtcctccga tagactgcgt cgcccgggta cccgtgtatc caataaaccc tcttgcagtt
     3721 gcatccgact tgtggtctcg ctgttccttg ggagggtctc ctctgagtga ttgactaccc
     3781 gtcagcgggg gtctttcatg ggtaacagtt tcttgaagtt ggagaacaac attctgaggg
     3841 taggagtcga atattaagta atcctgactc aattagccac tgttttgaat ccacatactc
     3901 caatactcct gaaatagttc attatggaca gcgcagaaag agctggggag aattgtgaaa
     3961 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     4021 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     4081 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     4141 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     4201 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     4261 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     4321 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     4381 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     4441 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     4501 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     4561 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     4621 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     4681 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     4741 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     4801 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     4861 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     4921 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     4981 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     5041 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     5101 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     5161 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     5221 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     5281 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     5341 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     5401 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     5461 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     5521 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     5581 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     5641 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     5701 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     5761 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     5821 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     5881 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     5941 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     6001 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     6061 gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt
     6121 aacctataaa aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg
     6181 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
     6241 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct
     6301 taactatgcg gcatcagagc agattgtact gagagtgcac catatgcggt gtgaaatacc
     6361 gcacagatgc gtaaggagaa aataccgcat caggcgccat tcgccattca ggctgcgcaa
     6421 ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg
     6481 atgtgctgca aggcgattaa gttgggtaac gccagggttt tcccagtcac gacgttgtaa
     6541 aacgacggcg caaggaatgg tgcatgcaag gagatggcgc ccaacagtcc cccggccacg
     6601 gggcctgcca ccatacccac gccgaaacaa gcgctcatga gcccgaagtg gcgagcccga
     6661 tcttccccat cggtgatgtc ggcgatatag gcgccagcaa ccgcacctgt ggcgccggtg
     6721 atgccggcca cgatgcgtcc ggcgtagagg cgattagtcc aatttgttaa agacaggata
     6781 tcagtggtcc aggctctagt tttgactcaa caatatcacc agctgaagcc tatagagtac
     6841 gagccataga taaaataaaa gattttattt agtctccaga aaaagggggg aatgaaagac
     6901 cccacctgta ggtttggcaa gctagcttaa gtaacgccat tttgcaaggc atggaaaata
     6961 cataactgag aatagagaag ttcagatcaa ggttaggaac agagagacag cagaatatgg
     7021 gccaaacagg atatctgtgg taagcagttc ctgccccggc tcagggccaa gaacagatgg
     7081 tccccagatg cggtcccgcc ctcagcagtt tctagagaac catcagatgt ttccagggtg
     7141 ccccaaggac ctgaaatgac cctgtgcctt atttgaacta accaatcagt tcgcttctcg
     7201 cttctgttcg cgcgcttctg ctccccgagc tcaataaaag agcccacaac ccctcactcg
     7261 gcgcgccagt cctccgatag ac
//
