LOCUS       sequence-322035-        7715 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 
            LTR promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7715)
  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 7715)
  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: pMY-CD3-P2A-CD90.2
FEATURES             Location/Qualifiers
     source          1..7715
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             22..615
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     CDS             1103..1519
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1487..1503
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     misc_feature    1529..1907
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     primer_bind     1845..1861
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     regulatory      1956..1965
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2523..2576
                     /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(5819..5836)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5990..6578)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6070..6089)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6742..7533,7534..7602))
                     /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     7365..7384
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7603..7707)
                     /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 attcccaata aagcctcttg ctgtttgcat
      541 ccgaatcgtg gactcgctga tccttgggag ggtctcctca gattgattga ctgcccacct
      601 cgggggtctt tcatttggag gttccaccga gatttggaga cccctgccca gggaccaccg
      661 acccccccgc cgggaggtaa gctggccagc ggtcgtttcg tgtctgtctc tgtctttgtg
      721 cgtgtttgtg ccggcatcta atgtttgcgc ctgcgtctgt actagttagc taactagctc
      781 tgtatctggc ggacccgtgg tggaactgac gagttcggaa cacccggccg caaccctggg
      841 agacgtccca gggacttcgg gggccgtttt tgtggcccga cctgagtcca aaaatcccga
      901 tcgttttgga ctctttggtg caccccccta ataggaggga tatgtggttc tggtaggaga
      961 cgagaaccta aaacagttcc cgcctccgtc tgaatttttg ctttcggttt gggaccgaag
     1021 ccgcgccgcg cgtcttgtct gctgcagcat cgttctgtgt tgtctctgtc tgactgtgtt
     1081 tctgtatttg tctgaaaatt agggccagac tgttaccact cccttaagtt tgaccttagg
     1141 tcactggaaa gatgtcgagc ggatcgctca caaccagtcg gtagatgtca agaagagacg
     1201 ttgggttacc ttctgctctg cagaatggcc aacctttaac gtcggatggc cgcgagacgg
     1261 cacctttaac cgagacctca tcacccaggt taagatcaag gtcttttcac ctggcccgca
     1321 tggacaccca gaccaggtcc cctacatcgt gacctgggaa gccttggctt ttgacccccc
     1381 tccctgggtc aagccctttg tacaccctaa gcctccgcct cctcttcctc catccgcccc
     1441 gtctctcccc cttgaacctc ctcgttcgac cccgcctcga tcctcccttt atccagccct
     1501 cactccttct ctaggcgccc ccatatggcc atatgagatc ttatatgggg cacccccgcc
     1561 ccttgtaaac ttccctgacc ctgacatgac aagagttact aacagcccct ctctccaagc
     1621 tcacttacag gctctctact tagtccagca cgaagtctgg agacctctgg cggcagccta
     1681 ccaagaacaa ctggaccgac cggccggtgg tacctcaccc ttaccgagtc ggcgacacag
     1741 tgtgggtccg ccgacaccag actaagaacc tagaacctcg ctggaaagga ccttacacag
     1801 tcctgctgac cacccccacc gccctcaaag tagacggcat cgcagcttgg atacacgccg
     1861 cccacgtgaa ggctgccgac cccgggggtg gaccatcctc tagactgccg gatctagcta
     1921 gttaattaac tccgtggatc cggtcgtgcg gccgcgccac catggaacag agaaaaggcc
     1981 tggccggcct gttcctggtt atcagtctgc tgcagggcac agtggcccag accaacaagg
     2041 ctaagaacct ggtgcaggtg gacggctcta gaggcgacgg atctgtgctg ctgacatgtg
     2101 gcctgaccga caagaccatc aagtggctga aggacggctc catcatcagc cctctgaacg
     2161 ccaccaagaa cacctggaac ctgggcaaca acgccaagga ccccagaggc acctatcagt
     2221 gccagggcgc caaagagaca agcaaccctc tgcaggtcta ctacagaatg tgcgagaact
     2281 gcatcgagct gaacatcggc accatcagcg gcttcatctt cgccgaagtg atcagcatct
     2341 tctttctggc cctgggcgtg tacctgatcg ctggacaaga tggcgtgcgg cagagcagag
     2401 ccagcgataa gcagacactg ctgcagaacg agcagctgta ccagcctctg aaggacagag
     2461 agtacgacca gtacagccac ctccagggca accagctgcg gaagaaggga tctggccaat
     2521 tggaaggcag aggctctctt cttacatgcg gcgacgtcga ggaaaaccca ggacctatgg
     2581 aacactctgg catcctggct agcctgatcc tgattgccgt tctgcctcaa ggcagcccct
     2641 tcaagatcca agtgaccgag tacgaggaca aggtgttcgt gacctgcaac accagcgtga
     2701 tgcacctgga tggcaccgtg gaaggatggt tcgccaagaa caagaccctg aacctcggca
     2761 agggcgtgct ggaccctaga ggcatctacc tgtgtaacgg cacagagcag ctggccaagg
     2821 tggtgtctag tgtgcaggtc cactatcgga tgtgtcagaa ctgcgtggaa ctggacagcg
     2881 gcacaatggc cggcgtgatc ttcatcgacc tgatcgctac cctgctgctg gcactgggag
     2941 tgtattgctt cgctggccac gagacaggca gacctagcgg agctgctgaa gttcaggccc
     3001 tgctgaagaa tgaacagctc tatcagcccc tgcgcgacag agaggatacc cagtactcta
     3061 gactcggcgg caactggccc agaaacaaga aatctggaag cggcacgcgt gtcagacaga
     3121 ccctgaactt cgatctgctt agactggccg gggacgtcga gtctaatcca ggaccaatgc
     3181 ggtggaacac cttctggggc atcctgtgtc tgtctctgct ggctgtgggc acctgtcagg
     3241 atgacgctga gaacatcgag tataaggtgt ccatctccgg caccagcgtc gagctgactt
     3301 gtcctctgga ctccgacgag aacctgaagt gggagaagaa cggccaagag ctgcctcaga
     3361 agcacgacaa gcacctggtg ctgcaggact tcagcgaggt ggaagatagc ggctactacg
     3421 tgtgctacac ccctgccagc aacaagaaca catacctgta cctgaaggct cgcgtgtgcg
     3481 agtactgtgt cgaggtggac ctgacagccg tggctatcat catcatcgtg gacatctgca
     3541 tcaccctggg cctgctgatg gtcatctact actggtccaa gaaccggaag gccaaggcca
     3601 agcctgtgac aagaggaacc ggcgctggaa gcagaccaag aggccagaac aaagaaagac
     3661 ctcctcctgt gcctaatcct gactacgagc ccatccggaa gggccagaga gatctgtact
     3721 ctggcctgaa ccagagggcc gtgggttctg gcgcatgcca gtgtaccaac tatgctctcc
     3781 tgagactcgc aggcgacgtt gagagtaatc cagggcctat gaagtggaaa gtgtctgtgc
     3841 tggcctgcat cctgcatgtt cgattccctg gcgctgaggc ccagtctttt ggactgctgg
     3901 accccaagct gtgctacctg ctggacggca ttctgtttat ttatggcgtg atcatcaccg
     3961 ctctgtacct gcgggccaag ttcagcagaa gcgctgagac agctgccaat ctgcaggacc
     4021 ctaaccagct gtacaacgag ctgaatctgg ggcgcagaga agagtacgat gtgctggaaa
     4081 agaagagagc cagagatccc gagatgggcg gcaaacagca gagaaggcgg aatcctcaag
     4141 aaggcgtgta caacgccctg cagaaagata agatggccga ggcctacagc gagatcggca
     4201 caaagggcga acgcagaaga ggcaagggac acgatggact gtaccagggc ctgtccacag
     4261 ccacaaagga cacatacgat gccctgcaca tgcagacact ggcccctaga ggcagcggcg
     4321 tcgacgccac aaacttcagc ctgctgagac aggctggcga cgtggaagag aatcctggac
     4381 ctatgaaccc cgccatctct gtggccctgc tgctgtctgt gctgcaggtg tccagaggcc
     4441 agaaagtgac cagcctgacc gcctgcctcg tgaaccagaa cctgagactg gactgcagac
     4501 acgagaacaa caccaaggac aacagcatcc agcacgagtt ctccctgacc cgcgagaaga
     4561 gaaaacacgt gctgagcggc accctgggca tccccgagca cacctacaga tccagagtga
     4621 ccctgagcaa ccagccctac atcaaggtgc tgaccctggc caacttcacc acaaaggacg
     4681 agggcgacta cttctgcgaa ctgcaggtgt caggcgccaa ccccatgagc agcaacaaga
     4741 gcatcagcgt gtacagagac aagctcgtga agtgcggcgg catcagcctg ctggtgcaga
     4801 acacctcttg gatgctgttg ctgctgctga gcctgagcct gctgcaggcc ctggacttca
     4861 tctctctgta gcctgcaggt ttctgaagct tgagtacatc gattgcattg ggtcgatcga
     4921 ccgggccgcc agcacagtgg tcgatcgagg gggggcccgc gattagtcca atttgttaaa
     4981 gacaggatat cagtggtcca ggctctagtt ttgactcaac aatatcacca gctgaagcct
     5041 atagagtacg agccatagat agaataaaag attttattta gtctccagaa aaagggggga
     5101 atgaaagacc ccacctgtag gtttggcaag ctagcttaag taacgccatt ttgcaaggca
     5161 tggaaaatac ataactgaga atagagaagt tcagatcaag gttaggaaca gagagacagc
     5221 agaatatggg ccaaacagga tatctgtggt aagcagttcc tgccccggct cagggccaag
     5281 aacagttgga acagcagaat atgggccaaa caggatatct gtggtaagca gttcctgccc
     5341 cggctcaggg ccaagaacag atggtcccca gatgcggtcc cgccctcagc agtttctaga
     5401 gaaccatcag atgtttccag ggtgccccaa ggacctgaaa tgaccctgtg ccttatttga
     5461 actaaccaat cagttcgctt ctcgcttctg ttcgcgcgct tctgctcccc gagctcaata
     5521 aaagagccca caacccctca ctcggcgcgc cagtcctccg atagactgcg tcgcccgggt
     5581 acccgtgtat ccaataaacc ctcttgcagt tgcatccgac ttgtggtctc gctgttcctt
     5641 gggagggtct cctctgagtg attgactacc cgtcagcggg ggtctttcac atgcagcatg
     5701 tatcaaaatt aatttggttt tttttcttaa gtatttacat taaatggcca tagttgcatt
     5761 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     5821 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     5881 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     5941 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     6001 tccgcacccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     6061 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     6121 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     6181 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     6241 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     6301 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     6361 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     6421 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     6481 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     6541 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     6601 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     6661 caaaaaggat cttcacctag atccttttgc ggccggccgc aaatcaatct aaagtatata
     6721 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     6781 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     6841 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     6901 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     6961 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     7021 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     7081 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     7141 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     7201 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     7261 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     7321 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     7381 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     7441 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     7501 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     7561 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     7621 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     7681 gaaaaataaa caaatagggg ttccgcgcac atttc
//
