LOCUS       sequence-322032-        8165 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Retroviral vector to overexpress the murine CD3 (TCR) components CD3
            gamma, CD delta, CD epsilon and TCR zeta and surface marker CD90.2, 
            separated by T2; F2A; E2A; P2A respectively, under control of the 
            CAG promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8165)
  AUTHORS   van der Donk LEH, van der Spek J, van Duivenvoorde T, Ten Brink MS, 
            Geijtenbeek TBH, Kuijl CP, van Heijst JWJ, Ates LS
  TITLE     An optimized retroviral toolbox for overexpression and genetic 
            perturbation of primary lymphocytes.
  JOURNAL   Biol Open. 2022 Feb 15;11(2):bio059032. doi: 10.1242/bio.059032. 
            Epub 2022 Mar 1.
  PUBMED    35229875
REFERENCE   2  (bases 1 to 8165)
  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: "Biol 
            Open."; date: "2022-02-15"; volume: "11(2):bio059032. doi"; pages: "
            10.1242/bio.059032. Epub 2022 Mar 1"
COMMENT     Sequence Label: pMX-CAG-CD3-P2A-CD90.2
FEATURES             Location/Qualifiers
     source          1..8165
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             22..615
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     misc_feature    678..1035
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1094..1510
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1440..1462
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1478..1494
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     misc_feature    1520..1898
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     primer_bind     1836..1852
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     enhancer        1923..2302
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2305..2580
                     /label=chicken beta-actin promoter
     regulatory      2801..2810
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3368..3421
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     primer_bind     complement(6269..6286)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6440..7028)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6520..6539)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7192..7983,7984..8052))
                     /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     7815..7834
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8053..8157)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cccgaaaagt gccacctgca taatgaaaga ccccacctgt aggtttggca agctagctta
       61 agtaacgcca ttttgcaagg catggaaaaa tacataactg agaatagaaa agttcagatc
      121 aaggtcagga acagatggaa cagctgaata tgggccaaac aggatatctg tggtaagcag
      181 ttcctgcccc ggctcagggc caagaacaga tggaacagct gaatatgggc caaacaggat
      241 atctgtggta agcagttcct gccccggctc agggccaaga acagatggtc cccagatgcg
      301 gtccagccct cagcagtttc tagagaacca tcagatgttt ccagggtgcc ccaaggacct
      361 gaaatgaccc tgtgccttat ttgaactaac caatcagttc gcttctcgct tctgttcgcg
      421 cgcttctgct ccccgagctc aataaaagag cccacaaccc ctcactcggc gcgccagtcc
      481 tccgattgac tgagtcgccc gggtacccgt gtatccaata aaccctcttg cagttgcatc
      541 cgacttgtgg tctcgctgtt ccttgggagg gtctcctctg agtgattgac tacccgtcag
      601 cgggggtctt tcatttgggg gctcgtccgg gatcgggaga cccctgccca gggaccaccg
      661 acccaccacc gggaggtaag ctggccagca acttatctgt gtctgtccga ttgtctagtg
      721 tctatgactg attttatgcg cctgcgtcgg tactagttag ctaactagct ctgtatctgg
      781 cggacccgtg gtggaactga cgagttcgga acacccggcc gcaaccctgg gagacgtccc
      841 agggacttcg ggggccgttt ttgtggcccg acctgagtcc aaaaatcccg atcgttttgg
      901 actctttggt gcacccccct aataggaggg atatgtggtt ctggtaggag acgagaacct
      961 aaaacagttc ccgcctccgt ctgaattttt gctttcggtt tgggaccgaa gccgcgccgc
     1021 gcgtcttgtc tgctgcagca tcgttctgtg ttgtctctgt ctgactgtgt ttctgtattt
     1081 gtctgaaaat tagggccaga ctgttaccac tcccttaagt ttgaccttag gtcactggaa
     1141 agatgtcgag cggatcgctc acaaccagtc ggtagatgtc aagaagagac gttgggttac
     1201 cttctgctct gcagaatggc caacctttaa cgtcggatgg ccgcgagacg gcacctttaa
     1261 ccgagacctc atcacccagg ttaagatcaa ggtcttttca cctggcccgc atggacaccc
     1321 agaccaggtc ccctacatcg tgacctggga agccttggct tttgaccccc ctccctgggt
     1381 caagcccttt gtacacccta agcctccgcc tcctcttcct ccatccgccc cgtctctccc
     1441 ccttgaacct cctcgttcga ccccgcctcg atcctccctt tatccagccc tcactccttc
     1501 tctaggcgcc cccatatggc catatgagat cttatatggg gcacccccgc cccttgtaaa
     1561 cttccctgac cctgacatga caagagttac taacagcccc tctctccaag ctcacttaca
     1621 ggctctctac ttagtccagc acgaagtctg gagacctctg gcggcagcct accaagaaca
     1681 actggaccga ccggccggtg gtacctcacc cttaccgagt cggcgacaca gtgtgggtcc
     1741 gccgacacca gactaagaac ctagaacctc gctggaaagg accttacaca gtcctgctga
     1801 ccacccccac cgccctcaaa gtagacggca tcgcagcttg gatacacgcc gcccacgtga
     1861 aggctgccga ccccgggggt ggaccatcct ctagactgcc ggatctagct agttaattaa
     1921 tcgacattga ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag
     1981 cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
     2041 caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg
     2101 gactttccat tgacgtcaat gggtggacta tttacggtaa actgcccact tggcagtaca
     2161 tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc
     2221 ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt
     2281 attagtcatc gctattacca tgggtcgagg tgagccccac gttctgcttc actctcccca
     2341 tctccccccc ctccccaccc ccaattttgt atttatttat tttttaatta ttttgtgcag
     2401 cgatgggggc gggggggggg gggncgcgcg ccaggcgggg cggggcgggg cgaggggcgg
     2461 ggcggggcga ggcggagagg tgcggcggca gccaatcaga gcggcgcgct ccgaaagttt
     2521 ccttttatgg cgaggcggcg gcggcggcgg ccctataaaa agcgaagcgc gcggcgggcg
     2581 ggagtcgctg cgttgccttc gccccgtgcc ccgctccgcg ccgcctcgcg ccgcccgccc
     2641 cggctctgac tgaccgcgtt actcccacag gtgagcgggc gggacggccc ttctcctccg
     2701 ggctgtaatt agcgcttggt ttaatgacgg ctcgtttctt ttctgtggct gcgtgaaagc
     2761 cttaaagggc tccgggaggg ggatccggtc gtgcggccgc gccaccatgg aacagagaaa
     2821 aggcctggcc ggcctgttcc tggttatcag tctgctgcag ggcacagtgg cccagaccaa
     2881 caaggctaag aacctggtgc aggtggacgg ctctagaggc gacggatctg tgctgctgac
     2941 atgtggcctg accgacaaga ccatcaagtg gctgaaggac ggctccatca tcagccctct
     3001 gaacgccacc aagaacacct ggaacctggg caacaacgcc aaggacccca gaggcaccta
     3061 tcagtgccag ggcgccaaag agacaagcaa ccctctgcag gtctactaca gaatgtgcga
     3121 gaactgcatc gagctgaaca tcggcaccat cagcggcttc atcttcgccg aagtgatcag
     3181 catcttcttt ctggccctgg gcgtgtacct gatcgctgga caagatggcg tgcggcagag
     3241 cagagccagc gataagcaga cactgctgca gaacgagcag ctgtaccagc ctctgaagga
     3301 cagagagtac gaccagtaca gccacctcca gggcaaccag ctgcggaaga agggatctgg
     3361 ccaattggaa ggcagaggct ctcttcttac atgcggcgac gtcgaggaaa acccaggacc
     3421 tatggaacac tctggcatcc tggctagcct gatcctgatt gccgttctgc ctcaaggcag
     3481 ccccttcaag atccaagtga ccgagtacga ggacaaggtg ttcgtgacct gcaacaccag
     3541 cgtgatgcac ctggatggca ccgtggaagg atggttcgcc aagaacaaga ccctgaacct
     3601 cggcaagggc gtgctggacc ctagaggcat ctacctgtgt aacggcacag agcagctggc
     3661 caaggtggtg tctagtgtgc aggtccacta tcggatgtgt cagaactgcg tggaactgga
     3721 cagcggcaca atggccggcg tgatcttcat cgacctgatc gctaccctgc tgctggcact
     3781 gggagtgtat tgcttcgctg gccacgagac aggcagacct agcggagctg ctgaagttca
     3841 ggccctgctg aagaatgaac agctctatca gcccctgcgc gacagagagg atacccagta
     3901 ctctagactc ggcggcaact ggcccagaaa caagaaatct ggaagcggca cgcgtgtcag
     3961 acagaccctg aacttcgatc tgcttagact ggccggggac gtcgagtcta atccaggacc
     4021 aatgcggtgg aacaccttct ggggcatcct gtgtctgtct ctgctggctg tgggcacctg
     4081 tcaggatgac gctgagaaca tcgagtataa ggtgtccatc tccggcacca gcgtcgagct
     4141 gacttgtcct ctggactccg acgagaacct gaagtgggag aagaacggcc aagagctgcc
     4201 tcagaagcac gacaagcacc tggtgctgca ggacttcagc gaggtggaag atagcggcta
     4261 ctacgtgtgc tacacccctg ccagcaacaa gaacacatac ctgtacctga aggctcgcgt
     4321 gtgcgagtac tgtgtcgagg tggacctgac agccgtggct atcatcatca tcgtggacat
     4381 ctgcatcacc ctgggcctgc tgatggtcat ctactactgg tccaagaacc ggaaggccaa
     4441 ggccaagcct gtgacaagag gaaccggcgc tggaagcaga ccaagaggcc agaacaaaga
     4501 aagacctcct cctgtgccta atcctgacta cgagcccatc cggaagggcc agagagatct
     4561 gtactctggc ctgaaccaga gggccgtggg ttctggcgca tgccagtgta ccaactatgc
     4621 tctcctgaga ctcgcaggcg acgttgagag taatccaggg cctatgaagt ggaaagtgtc
     4681 tgtgctggcc tgcatcctgc atgttcgatt ccctggcgct gaggcccagt cttttggact
     4741 gctggacccc aagctgtgct acctgctgga cggcattctg tttatttatg gcgtgatcat
     4801 caccgctctg tacctgcggg ccaagttcag cagaagcgct gagacagctg ccaatctgca
     4861 ggaccctaac cagctgtaca acgagctgaa tctggggcgc agagaagagt acgatgtgct
     4921 ggaaaagaag agagccagag atcccgagat gggcggcaaa cagcagagaa ggcggaatcc
     4981 tcaagaaggc gtgtacaacg ccctgcagaa agataagatg gccgaggcct acagcgagat
     5041 cggcacaaag ggcgaacgca gaagaggcaa gggacacgat ggactgtacc agggcctgtc
     5101 cacagccaca aaggacacat acgatgccct gcacatgcag acactggccc ctagaggcag
     5161 cggcgtcgac gccacaaact tcagcctgct gagacaggct ggcgacgtgg aagagaatcc
     5221 tggacctatg aaccccgcca tctctgtggc cctgctgctg tctgtgctgc aggtgtccag
     5281 aggccagaaa gtgaccagcc tgaccgcctg cctcgtgaac cagaacctga gactggactg
     5341 cagacacgag aacaacacca aggacaacag catccagcac gagttctccc tgacccgcga
     5401 gaagagaaaa cacgtgctga gcggcaccct gggcatcccc gagcacacct acagatccag
     5461 agtgaccctg agcaaccagc cctacatcaa ggtgctgacc ctggccaact tcaccacaaa
     5521 ggacgagggc gactacttct gcgaactgca ggtgtcaggc gccaacccca tgagcagcaa
     5581 caagagcatc agcgtgtaca gagacaagct cgtgaagtgc ggcggcatca gcctgctggt
     5641 gcagaacacc tcttggatgc tgttgctgct gctgagcctg agcctgctgc aggccctgga
     5701 cttcatctct ctgtagcctg caggtttctg aagcttgagt acatcgataa aataaaagat
     5761 tttatttagt ctccagaaaa aggggggaat gaaagacccc acctgtaggt ttggcaagct
     5821 agctagagaa ccatcagatg tttccagggt gccccaagga cctgaaatga ccctgtgcct
     5881 tatttgaact aaccaatcag ttcgcttctc gcttctgttc gcgcgcttct gctccccgag
     5941 ctcaataaaa gagcccacaa cccctcactc ggcgcgccag tcctccgata gactgcgtcg
     6001 cccgggtacc cgtgtatcca ataaaccctc ttgcagttgc atccgacttg tggtctcgct
     6061 gttccttggg agggtctcct ctgagtgatt gactacccgt cagcgggggt ctttcatggg
     6121 taacagtttc ttgaagtgtc gagcagcatg tatcaaaatt aatttggttt tttttcttaa
     6181 gtatttacat taaatggcca tagttgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     6241 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     6301 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     6361 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     6421 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     6481 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     6541 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     6601 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     6661 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     6721 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     6781 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     6841 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     6901 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     6961 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     7021 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     7081 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttgc
     7141 ggccggccgc aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     7201 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     7261 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     7321 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     7381 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     7441 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     7501 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     7561 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     7621 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     7681 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     7741 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     7801 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     7861 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     7921 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     7981 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     8041 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     8101 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     8161 atttc
//
