LOCUS       sequence-343612-        7337 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Generation of TVA-expressing stable cell lines, resistant to the 
            antibiotic Puromycin.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7337)
  AUTHORS   Sumser A, Joesch M, Jonas P, Ben-Simon Y
  TITLE     Fast, high-throughput production of improved rabies viral vectors 
            for specific, efficient and versatile transsynaptic retrograde 
            labeling.
  JOURNAL   Elife. 2022 Aug 30;11:e79848. doi: 10.7554/eLife.79848.
  PUBMED    36040301
REFERENCE   2  (bases 1 to 7337)
  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: "Elife."; 
            date: "2022-08-30"; volume: "11:e79848. doi"; pages: " 
            10.7554/eLife.79848"
COMMENT     Sequence Label: MMLV-CAG-TVA-IRES-Puro
FEATURES             Location/Qualifiers
     source          1..7337
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        17..396
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        397..600
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     551..571
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             601..776
                     /label=5' LTR (truncated)
                     /note="truncated long terminal repeat from Moloney murine 
                     sarcoma virus"
     misc_feature    839..1038
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1239..1655
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     1585..1607
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     1623..1639
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     primer_bind     2517..2539
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     2601..2620
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      2653..2662
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     misc_feature    join(3123..3697,3698..3700,3701..3709)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3290..3307)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3517..3536
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             3729..4328
                     /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(3729..3748)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4225..4245
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             4824..5293
                     /note="long terminal repeat from Moloney murine leukemia 
                     virus"
     primer_bind     complement(5423..5440)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5594..6182)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5674..5693)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6342..7133,7134..7202))
                     /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     6965..6984
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 acattaccgc catgttgaca ttgattattg actagttatt aatagtaatc aattacgggg
       61 tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg
      121 cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata
      181 gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc
      241 cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac
      301 ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg
      361 cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc
      421 aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc
      481 aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc
      541 gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct
      601 caataaaaga gcccacaacc cctcactcgg ggcgccagtc ctccgattga ctgagtcgcc
      661 cgggtacccg tattcccaat aaagcctctt gctgtttgca tccgaatcgt ggtctcgctg
      721 ttccttggga gggtctcctc tgagtgattg actacccacg acgggggtct ttcatttggg
      781 ggctcgtccg ggatttggag acccctgccc agggaccacc gacccaccac cgggaggtaa
      841 gctggccagc aacttatctg tgtctgtccg attgtctagt gtctatgttt gatgttatgc
      901 gcctgcgtct gtactagtta gctaactagc tctgtatctg gcggacccgt ggtggaactg
      961 acgagttctg aacacccggc cgcaaccctg ggagacgtcc cagggacttt gggggccgtt
     1021 tttgtggccc gacctgagga agggagtcga tgtggaatcc gaccccgtca ggatatgtgg
     1081 ttctggtagg agacgagaac ctaaaacagt tcccgcctcc gtctgaattt ttgctttcgg
     1141 tttggaaccg aagccgcgcg tcttgtctgc tgcagcgctg cagcatcgtt ctgtgttgtc
     1201 tctgtctgac tgtgtttctg tatttgtctg aaaattaggg ccagactgtt accactccct
     1261 taagtttgac cttaggtcac tggaaagatg tcgagcggat cgctcacaac cagtcggtag
     1321 atgtcaagaa gagacgttgg gttaccttct gctctgcaga atggccaacc tttaacgtcg
     1381 gatggccgcg agacggcacc tttaaccgag acctcatcac ccaggttaag atcaaggtct
     1441 tttcacctgg cccgcatgga cacccagacc aggtccccta catcgtgacc tgggaagcct
     1501 tggcttttga cccccctccc tgggtcaagc cctttgtaca ccctaagcct ccgcctcctc
     1561 ttcctccatc cgccccgtct ctcccccttg aacctcctcg ttcgaccccg cctcgatcct
     1621 ccctttatcc agccctcact ccttctctag gcgccggaat tgcagagagg gagtggccaa
     1681 ctccatcact aggggttcct tgtagttaat gattaacccg ccatgctact tatctacgta
     1741 gccatgctct aggaagagta ccattgacgt caataatgac gtatgttccc atagtaacgc
     1801 caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg
     1861 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat
     1921 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
     1981 tctacgtatt agtcatcgct attaccatgg tcgaggtgag ccccacgttc tgcttcactc
     2041 tccccatctc ccccccctcc ccacccccaa ttttgtattt atttattttt taattatttt
     2101 gtgcagcgat gggggcgggg cggggcgagg cggagaggtg cggcggcagc caatcagagc
     2161 ggcgcgctcc gaaagtttcc ttttatggcg aggcggcggc ggcggcggcc ctataaaaag
     2221 cgaagcgcgc gacgggcggg agtcgctgcg cgctgccttc gccccgtgcc ccgctccgcc
     2281 gccgcctcgc gccgcccgcc ccggctctga ctgaccgcgt tactcccaca ggtgagcggg
     2341 cgggacggcc cttctcctcc gggctgtaat tagcgcttgg tttaatgacg gcttgtttct
     2401 tttctgtggc tgcgtgaaag ccttgagggg ctccgggagg gccctttgtg cggggggagc
     2461 ggctcggggc tgtccgcggg gggacggctg ccttcggggg ggacggggca gggcggggtt
     2521 cggcttctgg cgtgtgaccg gcggctctag agcctctgct aaccatgttc atgccttctt
     2581 ctttttccta cagctcctgg gcaacgtgct ggttattgtg ctgtctcatc attttggcaa
     2641 agaattggat ccgccaccat ggcgcggctg ctgcccgcgc tgctgctgct gctgctgccc
     2701 ggtaacgtga ccggtaacgg gtccggtaac ggttctttgt cccgttgccc ccccggtcag
     2761 ttccgctgct cggagccgcc cggtgcccac ggggagtgtt acccgcagga ctggctgtgc
     2821 gacggacacc ccgactgcga cgacgggcgg gacgagtggg gctgcgggac cagcgcgatc
     2881 cccgcggtgc ccacggacaa cggcacagag gctcccactg cccctgctcc tggacgtgct
     2941 ctgccagcca ggaatcacgg ccgcatgtgg atgctgatca ctgcagtgct cctgtgctgc
     3001 ctggtagctg tgggtggtat cgctgcatgg gggaagtcca aagcaaaaag caggtctgac
     3061 atcttcagtc ttgaaagcgc atccaagggg ggtgggtgag aattccgccc cccccccccc
     3121 cccccctctc cctccccccc ccctaacgtt actggccgaa gccgcttgga ataaggccgg
     3181 tgtgcgtttg tctatatgtt attttccacc atattgccgt cttttggcaa tgtgagggcc
     3241 cggaaacctg gccctgtctt cttgacgagc attcctaggg gtctttcccc tctcgccaaa
     3301 ggaatgcaag gtctgttgaa tgtcgtgaag gaagcagttc ctctggaagc ttcttgaaga
     3361 caaacaacgt ctgtagcgac cctttgcagg cagcggaacc ccccacctgg cgacaggtgc
     3421 ctctgcggcc aaaagccacg tgtataagat acacctgcaa aggcggcaca accccagtgc
     3481 cacgttgtga gttggatagt tgtggaaaga gtcaaatggc tctcctcaag cgtattcaac
     3541 aaggggctga aggatgccca gaaggtaccc cattgtatgg gatctgatct ggggcctcgg
     3601 tgcacatgct ttacatgtgt ttagtcgagg ttaaaaaaac gtctaggccc cccgaaccac
     3661 ggggacgtgg ttttcctttg aaaaacacga tgataagctt gccacaaccc acaaggagac
     3721 gaccttccat gaccgagtac aagcccacgg tgcgcctcgc cacccgcgac gacgtccccc
     3781 gggccgtacg caccctcgcc gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg
     3841 acccggaccg ccacatcgag cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg
     3901 ggctcgacat cggcaaggtg tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca
     3961 cgccggagag cgtcgaagcg ggggcggtgt tcgccgagat cggcccgcgc atggccgagt
     4021 tgagcggttc ccggctggcc gcgcagcaac agatggaagg cctcctggcg ccgcaccggc
     4081 ccaaggagcc cgcgtggttc ctggccaccg tcggcgtctc gcccgaccac cagggcaagg
     4141 gtctgggcag cgccgtcgtg ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg
     4201 ccttcctgga gacctccgcg ccccgcaacc tccccttcta cgagcggctc ggcttcaccg
     4261 tcaccgccga cgtcgaggtg cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg
     4321 gtgcctgacg cccgccccac gacccgcagc gcccgaccga aaggagcgca cgaccccatg
     4381 gctccgaccg aagccacccg gggcggcccc gccgaccccg cacccgcccc cgaggcccac
     4441 cgactctagt cgagggctgc agcgctgcag aggccgagtg cagaactgct ccaaagggac
     4501 ctcaaggctt tccgagggac actaggctga ctccatcgag ccagtgtaga gataagctta
     4561 tcgattagtc caatttgtta aagacaggat atcagtggtc caggctctag ttttgactca
     4621 acaatatcac cagctgaagc ctatagagta cgagccatag ataaaataaa agattttatt
     4681 tagtctccag aaaaaggggg gaatgaaaga ccccacctgt aggtttggca agctagctta
     4741 agtaacgcca ttttgcaagg catggaaaaa tacataactg agaatagaga agttcagatc
     4801 aaggtcagga acagatggaa cagctgaata tgggccaaac aggatatctg tggtaagcag
     4861 ttcctgcccc ggctcagggc caagaacaga tggaacagct gaatatgggc caaacaggat
     4921 atctgtggta agcagttcct gccccggctc agggccaaga acagatggtc cccagatgcg
     4981 gtccagccct cagcagtttc tagagaacca tcagatgttt ccagggtgcc ccaaggacct
     5041 gaaatgaccc tgtgccttat ttgaactaac caatcagttc gcttctcgct tctgttcgcg
     5101 cgcttctgct ccccgagctc aataaaagag cccacaaccc ctcactcggg gcgccagtcc
     5161 tccgattgac tgagtcgccc gggtacccgt gtatccaata aaccctcttg cagttgcatc
     5221 cgacttgtgg tctcgctgtt ccttgggagg gtctcctctg agtgattgac tacccgtcag
     5281 cgggggtctt tcacatgcag catgtatcaa aattaatttg gttttttttc ttaagtattt
     5341 acattaaatg gccatagttg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     5401 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     5461 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     5521 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     5581 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     5641 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     5701 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     5761 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     5821 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     5881 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     5941 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     6001 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     6061 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     6121 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     6181 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     6241 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttgcggccgc
     6301 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     6361 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     6421 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     6481 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     6541 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     6601 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     6661 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     6721 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     6781 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     6841 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     6901 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     6961 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     7021 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     7081 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     7141 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     7201 atactcttcc tttttcaatg atccggccat tagccatatt attcattggt tatatagcat
     7261 aaatcaatat tggctattgg ccattgcata cgttgtatcc atatcataat atgtacattt
     7321 atattggctc atgtcca
//
