LOCUS       sequence-356198-        6620 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Retroviral expression vector containing the DivisionRecorder 
            cassette.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6620)
  AUTHORS   Bresser K, Kok L, Swain AC, King LA, Jacobs L, Weber TS, Perie L, 
            Duffy KR, de Boer RJ, Scheeren FA, Schumacher TN
  TITLE     Replicative history marks transcriptional and functional disparity 
            in the CD8(+) T cell memory pool.
  JOURNAL   Nat Immunol. 2022 May;23(5):791-801. doi: 
            10.1038/s41590-022-01171-9. Epub 2022 Apr 7.
  PUBMED    35393592
REFERENCE   2  (bases 1 to 6620)
  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: 
            "10.1038/s41590-022-01171-9"; journalName: "Nat Immunol"; date: 
            "2022-05"; volume: "23"; issue: "5"; pages: "791-801"
COMMENT     Sequence Label: pMX-DivisionRecorder
FEATURES             Location/Qualifiers
     source          1..6620
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     2..21
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(240..344)
                     /gene="bla"
                     /label=AmpR promoter
     LTR             374..967
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    1030..1387
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1452..1868
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1798..1820
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1836..1852
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     misc_feature    1878..2252
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     primer_bind     2190..2206
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     regulatory      2276..2285
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2282..2284,2285..2287,2288..2998)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2282..2284,2285..2287,2288..2998)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2588..2607)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2935..2956
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3008..3064
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     protein_bind    complement(3065..3098)
                     /label=loxP511
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATAC) (Shaw et al., 2021). loxP511 sites are 
                     compatible with each other, but incompatible with wild-type
                     loxP sites."
     CDS             4267..4296
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     protein_bind    complement(4311..4344)
                     /label=loxP511
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATAC) (Shaw et al., 2021). loxP511 sites are 
                     compatible with each other, but incompatible with wild-type
                     loxP sites."
     LTR             4537..5006
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(5076..5093)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5247..5835)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5327..5346)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5999..6620,1..170,171..239))
                     /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"
ORIGIN
        1 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
       61 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
      121 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
      181 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
      241 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
      301 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
      361 gtgccacctg cataatgaaa gaccccacct gtaggtttgg caagctagct taagtaacgc
      421 cattttgcaa ggcatggaaa aatacataac tgagaataga aaagttcaga tcaaggtcag
      481 gaacagatgg aacagctgaa tatgggccaa acaggatatc tgtggtaagc agttcctgcc
      541 ccggctcagg gccaagaaca gatggaacag ctgaatatgg gccaaacagg atatctgtgg
      601 taagcagttc ctgccccggc tcagggccaa gaacagatgg tccccagatg cggtccagcc
      661 ctcagcagtt tctagagaac catcagatgt ttccagggtg ccccaaggac ctgaaatgac
      721 cctgtgcctt atttgaacta accaatcagt tcgcttctcg cttctgttcg cgcgcttctg
      781 ctccccgagc tcaataaaag agcccacaac ccctcactcg gcgcgccagt cctccgattg
      841 actgagtcgc ccgggtaccc gtgtatccaa taaaccctct tgcagttgca tccgacttgt
      901 ggtctcgctg ttccttggga gggtctcctc tgagtgattg actacccgtc agcgggggtc
      961 tttcatttgg gggctcgtcc gggatcggga gacccctgcc cagggaccac cgacccacca
     1021 ccgggaggta agctggccag caacttatct gtgtctgtcc gattgtctag tgtctatgac
     1081 tgattttatg cgcctgcgtc ggtactagtt agctaactag ctctgtatct ggcggacccg
     1141 tggtggaact gacgagttcg gaacacccgg ccgcaaccct gggagacgtc ccagggactt
     1201 cgggggccgt ttttgtggcc cgacctgagt ccaaaaatcc cgatcgtttt ggactctttg
     1261 gtgcaccccc cttagaggag ggatatgtgg ttctggtagg agacgagaac ctaaaacagt
     1321 tcccgcctcc gtctgaattt ttgctttcgg tttgggaccg aagccgcgcc gcgcgtcttg
     1381 tctgctgcag catcgttctg tgttgtctct gtctgactgt gtttctgtat ttgtctgaaa
     1441 atatgggccc gggccagact gttaccactc ccttaagttt gaccttaggt cactggaaag
     1501 atgtcgagcg gatcgctcac aaccagtcgg tagatgtcaa gaagagacgt tgggttacct
     1561 tctgctctgc agaatggcca acctttaacg tcggatggcc gcgagacggc acctttaacc
     1621 gagacctcat cacccaggtt aagatcaagg tcttttcacc tggcccgcat ggacacccag
     1681 accaggtccc ctacatcgtg acctgggaag ccttggcttt tgacccccct ccctgggtca
     1741 agccctttgt acaccctaag cctccgcctc ctcttcctcc atccgccccg tctctccccc
     1801 ttgaacctcc tcgttcgacc ccgcctcgat cctcccttta tccagccctc actccttctc
     1861 taggcgcccc catatggcca tatgagatct tatatggggc acccccgccc cttgtaaact
     1921 tccctgaccc tgacatgaca agagttacta acagcccctc tctccaagct cacttacagg
     1981 ctctctactt agtccagcac gaagtctgga gacctctggc ggcagcctac caagaacaac
     2041 tggaccgacc ggtggtacct cacccttacc gagtcggcga cacagtgtgg gtccgccgac
     2101 accagactaa gaacctagaa cctcgctgga aaggacctta cacagtcctg ctgaccaccc
     2161 ccaccgccct caaagtagac ggcatcgcag cttggataca cgccgcccac gtgaaggctg
     2221 ccgaccccgg gggtggacca tcctctagac tgccggatct gattctagag ctagcgccac
     2281 catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga
     2341 cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta
     2401 cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac
     2461 cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg accacatgaa
     2521 gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt
     2581 cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct
     2641 ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca
     2701 caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca agcagaagaa
     2761 cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc
     2821 cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca
     2881 ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt
     2941 cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaaggg
     3001 ttccggagcc acgaacttct ctctgttaaa gcaagcagga gacgtggaag aaaaccccgg
     3061 tcccataact tcgtatagta tacattatac gaagttatca gaattcgggg gggggggggg
     3121 gggggggggg ctcgagggcc aatttactga ccgtacacca aaatttgcct gcattaccgg
     3181 tcgatgcaac gagtgatgag gttcgcaaga acctgtgtga ctgcttcagg gatcgccagg
     3241 cgttttctga gcatacctgg aaatgtcttc tgtccgtttg ccggtcgtgg gcggcatggt
     3301 gcaagttgaa taaccggaaa tggtttcccg cagaacctga agatgttcgc gattatcttc
     3361 tatatcttca ggcgcgcggt ctggcagtaa aaactatcca gcaacatttg ggccagctaa
     3421 acatgcttca tcgtcggtcc gggctgccac gaccaagtga cagcaatgct gtttcactgg
     3481 ttatgcggcg gatcagaaaa gaaaacgttg atgccggtga acgtgcaaaa caggctctag
     3541 cgttcgaacg cactgatttc gaccaggtaa gtagagggct gggctgggct gtggggggtg
     3601 tggggtgcgg gacttgggca tgtctgggag tccctctcac cacttttctt acctttctag
     3661 gttcgttcac tcatggaaaa tagcgatcgc tgccaggata tacgtaatct ggcatttctg
     3721 gggattgctt ataacaccct gttacgtata gccgaaattg ccaggatcag ggttaaagat
     3781 atctcacgta ctgacggtgg gagaatgtta atccatattg gcagaacgaa aacgctggtt
     3841 agcaccgcag gtgtagagaa ggcacttagc ctgggggtaa ctaaactggt cgagcgatgg
     3901 atttccgtct ctggtgtagc tgatgatccg aataactacc tgttttgccg ggtcagaaaa
     3961 aatggtgttg ccgcgccatc tgccaccagc cagctatcaa ctcgcgccct ggaagggatt
     4021 tttgaagcaa ctcatcgatt gatttacggc gctaaggatg actctggtca gagatacctg
     4081 gcctggtctg gacacagtgc ccgtgtcgga gccgcgcgag atatggcccg cgctggagtt
     4141 tcaataccgg agatcatgca agctggtggc tggaccaatg taaatattgt catgaactat
     4201 atccgtaacc tggatagtga aacaggggca atggtgcgcc tgctggaaga tggcgatgga
     4261 ccggtggaac aaaaacttat ttctgaagaa gatctgtgat agtaagtcgg ataacttcgt
     4321 atagtataca ttatacgaag ttatggatcc gcgccacgcg gccgcagtcg acgataaaat
     4381 aaaagatttt atttagtctc cagaaaaagg ggggaatgaa agaccccacc tgtaggtttg
     4441 gcaagctagc ttaagtaacg ccattttgca aggcatggaa aaatacataa ctgagaatag
     4501 agaagttcag atcaaggtca ggaacagatg gaacagctga atatgggcca aacaggatat
     4561 ctgtggtaag cagttcctgc cccggctcag ggccaagaac agatggaaca gctgaatatg
     4621 ggccaaacag gatatctgtg gtaagcagtt cctgccccgg ctcagggcca agaacagatg
     4681 gtccccagat gcggtccagc cctcagcagt ttctagagaa ccatcagatg tttccagggt
     4741 gccccaagga cctgaaatga ccctgtgcct tatttgaact aaccaatcag ttcgcttctc
     4801 gcttctgttc gcgcgcttct gctccccgag ctcaataaaa gagcccacaa cccctcactc
     4861 ggggcgccag tcctccgatt gactgagtcg cccgggtacc cgtgtatcca ataaaccctc
     4921 ttgcagttgc atccgacttg tggtctcgct gttccttggg agggtctcct ctgagtgatt
     4981 gactacccgt cagcgggggt ctttcacatg cwgcatkwat saawmkkmmw wcgcgcgggg
     5041 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     5101 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     5161 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     5221 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     5281 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     5341 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     5401 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     5461 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     5521 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     5581 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     5641 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     5701 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     5761 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     5821 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     5881 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     5941 cttttgcggc cggccgcaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     6001 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     6061 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     6121 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     6181 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     6241 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     6301 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     6361 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     6421 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     6481 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     6541 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     6601 cttgcccggc gtcaatacgg
//
