LOCUS       sequence-321432-        5880 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Retroviral (MSCV) expression of LSSmOrange fluorescent protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5880)
  AUTHORS   Landry LG, Mann SE, Anderson AM, Nakayama M
  TITLE     Multiplex T-cell Stimulation Assay Utilizing a T-cell Activation 
            Reporter-based Detection System.
  JOURNAL   Bio Protoc. 2021 Jan 20;11(2):e3883. doi: 10.21769/BioProtoc.3883. 
            eCollection 2021 Jan 20.
  PUBMED    33732772
REFERENCE   2  (bases 1 to 5880)
  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: "Bio 
            Protoc."; date: "2021-01-20"; volume: "11(2):e3883. doi"; pages: " 
            10.21769/BioProtoc.3883. eCollection 2021 Jan 20"
COMMENT     Sequence Label: pMSCVII-LO
FEATURES             Location/Qualifiers
     source          1..5880
                     /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      1414..1423
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1420..2130
                     /codon_start=1
                     /product="monomeric orange fluorescent protein with a large
                     Stokes shift (Shcherbakova et al., 2012)"
                     /label=LSSmOrange
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEENNMAIIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEG
                     FQTVKLKVTKGGPLPFAWDILSPQFTYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGMEASSERMYPEDGALK
                     GEDKLRLKLKDGGHYTSEVKTTYKAKKPVQLPGAYIVDIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     1833..1852
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     LTR             2275..2789
                     /label=3' LTR
                     /note="3' long terminal repeat from murine embryonic stem 
                     cell virus"
     protein_bind    2951..2967
                     /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(2975..2981,2982..2999,3000..3005))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3020..3041
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3158..3175)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3329..3917)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3409..3428)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4088..4879,4880..4948))
                     /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     4711..4730
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4949..5053)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5121..5139
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5177..5199)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5299..5318
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5512..5534
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5662..5681
                     /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 tcagccacca tggtgagcaa gggcgaggag
     1441 aataacatgg ccatcatcaa ggagttcatg cgcttcaagg tgcgcatgga gggctccgtg
     1501 aacggccacg agttcgagat cgagggcgag ggcgagggcc gcccctacga gggctttcag
     1561 accgttaagc tgaaggtgac caagggtggc cccctgccct tcgcctggga catcttgtcc
     1621 cctcagttca cctacggctc caaggcctac gtgaagcacc ccgccgacat ccccgactac
     1681 ctcaagctgt ccttccccga gggcttcaag tgggagcgcg tgatgaactt cgaggacggc
     1741 ggcgtggtga ccgtgactca ggactcctcc ctgcaggacg gcgagttcat ctacaaggtg
     1801 aagctgcgcg gcaccaactt cccctccgac ggccccgtaa tgcagaagaa gaccatgggc
     1861 atggaggcct cctccgagcg gatgtacccc gaggacggcg ccctgaaggg cgaggacaag
     1921 ctcaggctga agctgaagga cggcggccac tacacctccg aggtcaagac cacctacaag
     1981 gccaagaagc ccgtgcagtt gcccggcgcc tacatcgtcg acatcaagtt ggacatcacc
     2041 tcccacaacg aggactacac catcgtggaa cagtacgaac gcgccgaggg ccgccactcc
     2101 accggcggca tggacgagct gtacaagtaa ctcgagcctg caggcatgca agcttcaggt
     2161 agccggctaa cgttaacaac cggtacctct agaactatag ctagcatgcg caaatttaaa
     2221 gcgctgatat cgataaaata aaagatttta tttagtctcc agaaaaaggg gggaatgaaa
     2281 gaccccacct gtaggtttgg caagctagct taagtaacgc cattttgcaa ggcatggaaa
     2341 atacataact gagaatagag aagttcagat caaggttagg aacagagaga cagcagaata
     2401 tgggccaaac aggatatctg tggtaagcag ttcctgcccc ggctcagggc caagaacaga
     2461 tggtccccag atgcggtccc gccctcagca gtttctagag aaccatcaga tgtttccagg
     2521 gtgccccaag gacctgaaat gaccctgtgc cttatttgaa ctaaccaatc agttcgcttc
     2581 tcgcttctgt tcgcgcgctt ctgctccccg agctcaataa aagagcccac aacccctcac
     2641 tcggcgcgcc agtcctccga tagactgcgt cgcccgggta cccgtgtatc caataaaccc
     2701 tcttgcagtt gcatccgact tgtggtctcg ctgttccttg ggagggtctc ctctgagtga
     2761 ttgactaccc gtcagcgggg gtctttcatg ggtaacagtt tcttgaagtt ggagaacaac
     2821 attctgaggg taggagtcga atattaagta atcctgactc aattagccac tgttttgaat
     2881 ccacatactc caatactcct gaaatagttc attatggaca gcgcagaaag agctggggag
     2941 aattgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     3001 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     3061 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     3121 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     3181 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     3241 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     3301 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     3361 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     3421 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     3481 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     3541 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     3601 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     3661 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     3721 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     3781 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     3841 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     3901 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     3961 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     4021 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     4081 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     4141 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     4201 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     4261 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     4321 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     4381 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     4441 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     4501 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     4561 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     4621 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     4681 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     4741 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     4801 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     4861 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     4921 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     4981 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     5041 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta
     5101 tcatgacatt aacctataaa aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg
     5161 gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt
     5221 aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc
     5281 ggggctggct taactatgcg gcatcagagc agattgtact gagagtgcac catatgcggt
     5341 gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggcgccat tcgccattca
     5401 ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta cgccagctgg
     5461 cgaaaggggg atgtgctgca aggcgattaa gttgggtaac gccagggttt tcccagtcac
     5521 gacgttgtaa aacgacggcg caaggaatgg tgcatgcaag gagatggcgc ccaacagtcc
     5581 cccggccacg gggcctgcca ccatacccac gccgaaacaa gcgctcatga gcccgaagtg
     5641 gcgagcccga tcttccccat cggtgatgtc ggcgatatag gcgccagcaa ccgcacctgt
     5701 ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg cgattagtcc aatttgttaa
     5761 agacaggata tcagtggtcc aggctctagt tttgactcaa caatatcacc agctgaagcc
     5821 tatagagtac gagccataga taaaataaaa gattttattt agtctccaga aaaagggggg
//
