LOCUS       sequence-316612-        6238 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Retroviral vector to express cytosolic mTurquoise2 and an HA-tagged 
            puromycin selection marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6238)
  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 6238)
  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-mTurquoise2-P2A-PuroR-HA
FEATURES             Location/Qualifiers
     source          1..6238
                     /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..1903
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     primer_bind     1841..1857
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     regulatory      1964..1973
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1970..1972,1973..1975,1976..2686)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Goedhart et 
                     al., 2012)"
                     /label=mTurquoise2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYFSDNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2015..2036)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2276..2295)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2623..2644
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2759..3355
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(2759..2778)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     3255..3275
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             3365..3391
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(4342..4359)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4513..5101)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4593..4612)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5265..6056,6057..6125))
                     /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     5888..5907
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6126..6230)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cccgaaaagt gccacctgca taatgaaaga ccccacctgt aggtttggca agctagctta
       61 agtaacgcca ttttgcaagg catggaaaaa tacataactg agaatagaaa agttcagatc
      121 aaggtcagga acagatggaa cagctgaata tgggccaaag cggatatctg 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 ctcactcggg gcgccagtcc
      481 tccgattgac tgagtcgccc gggtacccgt attcccaata aagcctcttg ctgtttgcat
      541 ccgaatcgtg gactcgctga tccttgggag ggtctcctca gattgattga ctgcccacct
      601 cgggggtctt tcatttggag gttccaccga gatttggaga ccccagccca 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 cggtggtacc tcacccttac cgagtcggcg acacagtgtg
     1741 ggtccgccga caccagacta agaacctaga acctcgctgg aaaggacctt acacagtcct
     1801 gctgaccacc cccaccgccc tcaaagtaga cggcatcgca gcttggatac acgccgccca
     1861 cgtgaaggct gccgaccccg ggggtggacc atcctctaga ctgccggatc tagctagtta
     1921 attaactccg tggatccggt cgtgcggccg cacggaaacg cgtgccacca tggtgagcaa
     1981 gggcgaggag ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa
     2041 cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac
     2101 cctgaagttc atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac
     2161 cctgtcctgg ggcgtgcagt gcttcgcccg ctaccccgac cacatgaagc agcacgactt
     2221 cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga
     2281 cggcaactac aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat
     2341 cgagctgaag ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta
     2401 caactacttt agcgacaacg tctatatcac cgccgacaag cagaagaacg gcatcaaggc
     2461 caacttcaag atccgccaca acatcgagga cggcggcgtg cagctcgccg accactacca
     2521 gcagaacacc cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac
     2581 ccagtccaag ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt
     2641 cgtgaccgcc gccgggatca ctctcggcat ggacgagctg tacaagggca gcggcgtcga
     2701 cgccacaaac ttcagcctgc tgagacaggc tggcgacgtg gaagagaatc ctggacctat
     2761 gaccgagtac aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg
     2821 caccctcgcc gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg
     2881 ccacatcgag cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat
     2941 cggcaaggtg tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag
     3001 cgtcgaagcg ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc
     3061 ccggctggcc gcgcagcaac agatggaagg cctcctggcg ccgcaccggc ccaaggagcc
     3121 cgcgtggttc ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag
     3181 cgccgtcgtg ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga
     3241 gacctccgcg ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga
     3301 cgtcgaggtg cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgccggcag
     3361 cggctaccct tacgatgtgc ctgattacgc ctgacctgca ggtttctgaa gcttgagtac
     3421 atcgattgca ttgggtcgat cgaccgggcc gccagcacag tggtcgatcg agggggggcc
     3481 cgcgattagt ccaatttgtt aaagacagga tatcagtggt ccaggctcta gttttgactc
     3541 aacaatatca ccagctgaag cctatagagt acgagccata gatagaataa aagattttat
     3601 ttagtctcca gaaaaagggg ggaatgaaag accccacctg taggtttggc aagctagctt
     3661 aagtaacgcc attttgcaag gcatggaaaa tacataactg agaatagaga agttcagatc
     3721 aaggttagga acagagagac agcagaatat gggccaaaca ggatatctgt ggtaagcagt
     3781 tcctgccccg gctcagggcc aagaacagtt ggaacagcag aatatgggcc aaacaggata
     3841 tctgtggtaa gcagttcctg ccccggctca gggccaagaa cagatggtcc ccagatgcgg
     3901 tcccgccctc agcagtttct agagaaccat cagatgtttc cagggtgccc caaggacctg
     3961 aaatgaccct gtgccttatt tgaactaacc aatcagttcg cttctcgctt ctgttcgcgc
     4021 gcttctgctc cccgagctca ataaaagagc ccacaacccc tcactcggcg cgccagtcct
     4081 ccgatagact gcgtcgcccg ggtacccgtg tatccaataa accctcttgc agttgcatcc
     4141 gacttgtggt ctcgctgttc cttgggaggg tctcctctga gtgattgact acccgtcagc
     4201 gggggtcttt cacatgcagc atgtatcaaa attaatttgg ttttttttct taagtattta
     4261 cattaaatgg ccatagttgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     4321 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     4381 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     4441 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     4501 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     4561 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     4621 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     4681 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     4741 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     4801 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     4861 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     4921 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     4981 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     5041 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     5101 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     5161 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt tgcggccggc
     5221 cgcaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     5281 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     5341 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     5401 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     5461 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     5521 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     5581 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     5641 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     5701 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     5761 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     5821 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     5881 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     5941 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     6001 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     6061 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     6121 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     6181 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttc
//
