LOCUS       sequence-321690-        5750 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Retroviral vector for knockdown of murine CD247A (TCR zeta) and 
            expression of a CD90.2 surface marker under control of the murine 
            PGK promoter. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5750)
  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 5750)
  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-mPGK-CD90.2-Cd247A_miR
FEATURES             Location/Qualifiers
     source          1..5750
                     /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"
     promoter        1931..2430
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1966..1989)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2363..2382
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     ncRNA           2995..3116
                     /label=5' miR-30a
                     /note="sequence upstream of the 71-nt precursor of the 
                     human miR-30a microRNA (Zeng et al., 2002)"
                     /ncRNA_class="miRNA"
     ncRNA           3144..3158
                     /label=miR-30a loop
                     /note="loop from the the 71-nt precursor of the human 
                     miR-30a microRNA (Zeng et al., 2002)"
                     /ncRNA_class="miRNA"
     ncRNA           3215..3327
                     /label=3' miR-30a
                     /note="sequence downstream of the 71-nt precursor of the 
                     human miR-30a microRNA (Zeng et al., 2002)"
                     /ncRNA_class="miRNA"
     primer_bind     complement(3854..3871)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4025..4613)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4105..4124)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4777..5568,5569..5637))
                     /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     5400..5419
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5638..5742)
                     /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 aaattctacc gggtagggga ggcgcttttc ccaaggcagt ctggagcatg cgctttagca
     1981 gccccgctgg gcacttggcg ctacacaagt ggcctctggc ctcgcacaca ttccacatcc
     2041 accggtaggc gccaaccggc tccgttcttt ggtggcccct tcgcgccacc ttctactcct
     2101 cccctagtca ggaagttccc ccccgccccg cagctcgcgt cgtgcaggac gtgacaaatg
     2161 gaagtagcac gtctcactag tctcgtgcag atggacagca ccgctgagca atggaagcgg
     2221 gtaggccttt ggggcagcgg ccaatagcag ctttgctcct tcgctttctg ggctcagagg
     2281 ctgggaaggg gtgggtccgg gggcgggctc aggggcgggc tcaggggcgg ggcgggcgcc
     2341 cgaaggtcct ccggaggccc ggcattctgc acgcttcaaa agcgcacgtc tgccgcgctg
     2401 ttctcctctt cctcatctcc gggcctttcg acggatccgg tcgtgcggcc gcacggaaac
     2461 gcgtgccacc atgaaccccg ccatctctgt ggccctgctg ctgtctgtgc tgcaggtgtc
     2521 cagaggccag aaagtgacca gcctgaccgc ctgcctcgtg aaccagaacc tgagactgga
     2581 ctgcagacac gagaacaaca ccaaggacaa cagcatccag cacgagttct ccctgacccg
     2641 cgagaagaga aaacacgtgc tgagcggcac cctgggcatc cccgagcaca cctacagatc
     2701 cagagtgacc ctgagcaacc agccctacat caaggtgctg accctggcca acttcaccac
     2761 aaaggacgag ggcgactact tctgcgaact gcaggtgtca ggcgccaacc ccatgagcag
     2821 caacaagagc atcagcgtgt acagagacaa gctcgtgaag tgcggcggca tcagcctgct
     2881 ggtgcagaac acctcttgga tgctgttgct gctgctgagc ctgagcctgc tgcaggccct
     2941 ggacttcatc tctctgtagg catgccgcga ccctgcaggt ttctgaagct ttgtttgaat
     3001 gaggcttcag tactttacag aatcgttgcc tgcacatctt ggaaacactt gctgggatta
     3061 cttcttcagg ttaacccaac agaaggctcg agaaggtata ttgctgttga cagtgagcgc
     3121 cctcttcatc tacggagtca ttagtgaagc cacagatgta atgactccgt agatgaagag
     3181 gatgcctact gcctcggact tcaaggggct agaattcaag gggctacttt aggagcaatt
     3241 atcttgttta ctaaaactga ataccttgct atctctttga tacattttta caaagctgaa
     3301 ttaaaatggt ataaattaaa tcactttatc gataaaataa aagattttat ttagtctcca
     3361 gaaaaagggg ggaatgaaag accccacctg taggtttggc aagctagcta gagaaccatc
     3421 agatgtttcc agggtgcccc aaggacctga aatgaccctg tgccttattt gaactaacca
     3481 atcagttcgc ttctcgcttc tgttcgcgcg cttctgctcc ccgagctcaa taaaagagcc
     3541 cacaacccct cactcggcgc gccagtcctc cgatagactg cgtcgcccgg gtacccgtgt
     3601 atccaataaa ccctcttgca gttgcatccg acttgtggtc tcgctgttcc ttgggagggt
     3661 ctcctctgag tgattgacta cccgtcagcg ggggtctttc atgggtaaca gtttcttgaa
     3721 gtgtcgagca gcatgtatca aaattaattt ggtttttttt cttaagtatt tacattaaat
     3781 ggccatagtt gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     3841 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     3901 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     3961 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     4021 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     4081 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     4141 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     4201 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc
     4261 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     4321 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     4381 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     4441 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag
     4501 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     4561 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     4621 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     4681 tggtcatgag attatcaaaa aggatcttca cctagatcct tttgcggccg gccgcaaatc
     4741 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     4801 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     4861 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     4921 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     4981 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     5041 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     5101 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     5161 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     5221 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     5281 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     5341 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     5401 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     5461 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     5521 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     5581 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     5641 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     5701 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
//
