LOCUS       sequence-393494-        5694 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  puroR nls-mNG marked rapid RMCE integration vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5694)
  AUTHORS   Nonet ML
  TITLE     Rapid generation of C. elegans single-copy transgenes combining RMCE
            and drug selection.
  JOURNAL   Genetics. 2023 Apr 20:iyad072. doi: 10.1093/genetics/iyad072.
  PUBMED    37079426
REFERENCE   2  (bases 1 to 5694)
  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: "Genetics. 
            2023 Apr 20:iyad072. doi: 10.1093/genetics/iyad072."
COMMENT     Sequence Label: pPuroG1
FEATURES             Location/Qualifiers
     source          1..5694
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(365..384)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     963..983
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             complement(2929..2949)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     protein_bind    complement(2962..2995)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     protein_bind    3016..3049
                     /label=FRT3
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision with other FRT3 sites
                     (Bosch et al., 2015)"
     protein_bind    3147..3180
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     rep_origin      3219..3674
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3306..3325)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3516..3537
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3700..3804
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3805..3873,3874..4665)
                     /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     complement(4023..4042)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4836..5424
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5325..5344
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5578..5595
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 catttttgct ttcgtcgtaa atctacacac gcgtctcttc cgtgcgagag tccaagccag
       61 cagccaaatt cgttgactga gtattcaacg tttatacgtt gtcggcaacg agaaatagga
      121 aaatgcatcg ggaaatgttc ttttttcgat tttttccaag gttttgacaa attttaccac
      181 gaattttgct atgttttcaa ttaaaaaata tgttattcaa ctgtttctat gaggaaaata
      241 aggctttgca tgtaattttc ttattcagca taatttttaa ttaatttgaa ttttctgtcc
      301 taacgtttat tttgttttct tggttatgac tgatctgaaa ttaatttttg aattttaagg
      361 taatatgacc gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg tccccagggc
      421 cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca ccgtcgatcc
      481 ggaccgccac atcgagcggg tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct
      541 cgacatcggc aaggtgtggg tcggtaagtt taaacatata tatactaact aaccctgatt
      601 atttaaattt tcagcggacg acggcgccgc ggtggcggtc tggaccacgc cggagagcgt
      661 cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg gccgagttga gcggttcccg
      721 gctggccgcg cagcaacaga tggaaggcct cctggcgccg caccggccca aggagcccgc
      781 gtggttcctg gccaccgtcg gcgtctcgcc cgacgtaagt ttaaacatga ttttactaac
      841 taactaatct gatttaaatt ttcagcacca gggcaagggt ctgggcagcg ccgtcgtgct
      901 ccccggagtg gaggcggccg agcgcgccgg ggtgcccgcc ttcctggaaa cctccgcgcc
      961 ccgcaacctc cccttctacg agcggctcgg cttcaccgtc accgccgacg tcgaggtgcc
     1021 cgaaggaccg cgcacctggt gcatgacccg caagcccggt gcctgagtcc aattactctt
     1081 caacatccct acatgctctt tctccctgtg ctcccacccc ctatttttgt tattatcaaa
     1141 aaacttctct taatttcttt gttttttagc ttcttttaag tcacctctaa caatgaaatt
     1201 gtgtagattc aaaaatagaa ttaattcgta ataaaaagtc gaaaaaaatt gtgctccctc
     1261 cccccattaa taataattct atcccaaaat ctacacaatg ttctgtgtac acttcttatg
     1321 ttttttactt ctgataaatt tttttgaaac atcatagaaa aaaccgcaca caaaatacct
     1381 tatcatatgt tacgtttcag tttatgaccg caatttttag acctgttcga ggacaactgg
     1441 gaaactttta caaacttaca acatggttca gacaagttta aaaatttaag tttgagaatc
     1501 aacgggactt tttctttttt cgtttttttt aattgtgtat gctttgaaga ttttcaaatt
     1561 tattctagtt tcaataaaaa cagtaatata attgagtggg agtttttttg ctgtgatgtt
     1621 tttggtgtta tttacagtga tcttcttcta aatagaaaat caatggttta ttgatcaaaa
     1681 atattgagaa gtgtcgaatc aattgaagca ttgaaggaca aatttccact atcctacaaa
     1741 tccgatgcgt aaaaattatt aaacaaagtg aaaaaaatgc atttgaagaa aaaaaacaac
     1801 agagaaggtg ttgatgggct tttattgtga cacgttgaat gtgtatgtgt tactgtgcag
     1861 tggtaggttt gggggacgta ggggaagaga aacgggtaaa gagattgaaa gaaaagagag
     1921 aaacgagatg gaattagagc agttgatcta cttggcgagc ttcttggcgt tctcggagag
     1981 cttggttggg ttggccttac gacgacggga ggccttgtag agttcatcca ttcccatcac
     2041 atctgtgaag gccttttgcc actctttgaa gttcagctct gtctttgagt gcttgagttc
     2101 tgtctttcgg aacacgtaca ttggttgatt ctgaaaattt aaatcagatt agttagttag
     2161 taaaatcatg tttaaactta ccttgaggta gttggcggcc attggcttag cgaatgtgta
     2221 tgttgttcgg gctgtcgatc ggtatcgctt tccgtttcca gtagtgtacg accacttgaa
     2281 tgtcgagatg attgtcttat catttgggta tgtctttttc gatcggcacc aatcagcagc
     2341 tgtgagtgag tttgtcatca ctggtccatc agctgggaat cctgttccct taacttgagc
     2401 ctctccctga aaatttaaat atgtatggtt agttagtacc gaactgttta aacttacctt
     2461 gatgtgcgat ccctcgtatg tgtatcggta gttcactgtg agcgaggctc catcctcgaa
     2521 ttgcattgtt cggtgaactt ggtatcccga tccatccacc atggcagctt ggaatggtga
     2581 cattccatct gaaaatttaa atattactcg ttagttagta gaactcagtt taaacttacc
     2641 tgggtatggg aggtattggt ggaatccgta tccgatgtgt ggcacgagaa tccatggcga
     2701 gaattgcaga tctccctttg tcgacttgag gttgagttcc tcgtatccat cgtttgggtt
     2761 tccagttcct gaaaatttaa ataatcaggg ttagttagta tatatatgtt taaacttacc
     2821 ttgtccgacc atatcgaaat ccactccgtt gatcgatccg aagatgtgga gttcgtgtgt
     2881 agctgggagt gaagccatgt tatcctcttc tcccttcgac accatcgaca ccttacgctt
     2941 cttctttggg gatcccattt tgaagttcct atactttcta gagaatagga acttcaccga
     3001 tatcgaattc ctgcagaagt tcctattctt caaatagtat aggaacttcc cgggtaccga
     3061 agagcatgca acgcgtggtt acctgcaggc tcttcgccac tagttctaga gcggccgcca
     3121 ccgcggtgga gctccaattc gcccgtataa cttcgtataa tgtatgctat acgaagttat
     3181 ccatctagag cggccgccac cgcggtggag ctccgacgac gcgccctgta gcggcgcatt
     3241 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     3301 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     3361 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     3421 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     3481 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     3541 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     3601 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt
     3661 aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     3721 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     3781 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     3841 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     3901 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     3961 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     4021 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     4081 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     4141 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     4201 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     4261 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     4321 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     4381 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     4441 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     4501 tctggagccg gtgagcgtgg atctcgcggt atcattgcag cactggggcc agatggtaag
     4561 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     4621 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     4681 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     4741 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     4801 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     4861 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     4921 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     4981 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     5041 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     5101 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaatgggg
     5161 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     5221 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     5281 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     5341 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     5401 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     5461 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     5521 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     5581 gagtcagtga gcgaggaagc ggaattcgcc caatacgcaa accgcctctc cccgcgcgtt
     5641 ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggat
//
