LOCUS       sequence-42403-e        5880 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5880)
  AUTHORS   Semple JI, Biondini L, Lehner B
  TITLE     Generating transgenic nematodes by bombardment and antibiotic 
            selection.
  JOURNAL   Nat Methods. 2012 Jan 30;9(2):118-9. doi: 10.1038/nmeth.1864.
  PUBMED    22290182
REFERENCE   2  (bases 1 to 5880)
  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"; doi: "10.1038/nmeth"; 
            journalName: "Nat Methods"; date: "2012-01-30- 30"; volume: "9"; 
            issue: "2"; pages: "118-9"
COMMENT     Sequence Label: pBCN42-R4R3
FEATURES             Location/Qualifiers
     source          1..5880
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(782..801)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1380..1400
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     2061..2083
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2121..2139)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2207..2311
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2312..2380,2381..3172)
                     /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(2530..2549)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3343..3931
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3832..3851
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4057..4079
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4076..4092
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4076..4092
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     protein_bind    4112..4235
                     /gene="mutant version of attR"
                     /label=attR4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(4276..4581)
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     complement(4378..4397)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(4901..5584)
                     /codon_start=1
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGRKRR
                     VDPAY"
     primer_bind     5499..5518
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(join(5638..5644,5645..5662,5663..5668))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(5693..5816)
                     /gene="mutant version of attR"
                     /label=attR3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(5824..5841)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5824..5840)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5833..5855)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga
       61 tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg cgcctgatgc ggtattttct
      121 ccttacgcat ctgtgcggta tttcacaccg catatgccgg gcccccctcg aggggagaaa
      181 atgcggagag aagaacccag catcttaata aaattatata attttgaata aaatcaaatt
      241 aaaaataaga ttaattgaaa cacaacaata gaatgcagaa aattactcat aagaaataat
      301 ttattttgaa agaagttgtg tcgatttacg ataaaaacta cacgcaaacg aggctccgcg
      361 tttacaccgt ttggcaatgg agcgcgcttg ctattagatt ttaggccagc tcaacgaaag
      421 tttaatttat ttttcttctt tttttccatt tttttgagta aaatatcgaa ttttttgtat
      481 taattctgta aattttataa tttgcaacta ttttggcgta ttctgcttgt ttttatcagc
      541 gaatttttat gcatttcagt caaaagtttg agcaggaaaa atgacattta ggactttgta
      601 aaattgtatt cttaaatttg cttgaaaacc tacgtgtaca attgatttta ttgaattttt
      661 cttacgtttc ccgttgtatc ttattgtatt taactttcat cataaaataa tttatttcca
      721 ggtaaaatat ccgacgctct cgtggatccc cgggattggc tcagaaaaaa aggtaccgac
      781 catgaccgag tacaagccca cggtgcgcct cgccacccgc gacgacgtcc ccagggccgt
      841 acgcaccctc gccgccgcgt tcgccgacta ccccgccacg cgccacaccg tcgatccgga
      901 ccgccacatc gagcgggtca ccgagctgca agaactcttc ctcacgcgcg tcgggctcga
      961 catcggcaag gtgtgggtcg gtaagtttaa acatatatat actaactaac cctgattatt
     1021 taaattttca gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga
     1081 agcgggggcg gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct
     1141 ggccgcgcag caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg
     1201 gttcctggcc accgtcggcg tctcgcccga cgtaagttta aacatgattt tactaactaa
     1261 ctaatctgat ttaaattttc agcaccaggg caagggtctg ggcagcgccg tcgtgctccc
     1321 cggagtggag gcggccgagc gcgccggggt gcccgccttc ctggagacct ccgcgccccg
     1381 caacctcccc ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga
     1441 aggaccgcgc acctggtgca tgacccgcaa gcccggtgcc tgatgagcta gccggccata
     1501 caagtaatcc ggatgatcga cgccaacgtc gttgaatttt caaattttaa atactgaata
     1561 tttgtttttt ttcctattat ttatttattc tctttgtgtt ttttttcttg ctttctaaaa
     1621 aattaattca atccaaatct aaacattttt ttttctcttt ccgtctccca attcgtattc
     1681 cgctcctctc atctgaacac aatgtgcaag tttatttatc ttctcgcttt catttcatta
     1741 ggacgtgggg ggaattggtg gaagggggaa acacacaaaa ggatgatgga aatgaaataa
     1801 ggacacacaa tatgcaacaa cattcaattc agaaatatgg aggaaggttt aaaagaaaac
     1861 ataaaaatat atagaggagg aaggaaaact agtccaagat ctcagtggtc ctaggtgagc
     1921 tcccatatgg tgcactctca gtgcaatctg ctctgatgcc gcatagttaa gccagccccg
     1981 acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta
     2041 cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc
     2101 gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat
     2161 aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     2221 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     2281 aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     2341 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     2401 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     2461 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt
     2521 taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg
     2581 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
     2641 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
     2701 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
     2761 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
     2821 cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa
     2881 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
     2941 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
     3001 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga
     3061 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
     3121 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
     3181 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     3241 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     3301 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     3361 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     3421 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     3481 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     3541 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     3601 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     3661 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     3721 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     3781 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     3841 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     3901 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     3961 cctggtcttt tgctggcctt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     4021 gataaccgta ttaccgcctt tgagtgagct gataccagcg gataacaatt tcacacagga
     4081 aacagctatg accatgatta cgccaagcta tcaactttgt atagaaaagt tgaacgagaa
     4141 acgtaaaatg atataaatat caatatatta aattagattt tgcataaaaa acagactaca
     4201 taatactgta aaacacaaca tatccagtca ctatggtcga cctgcagact ggctgtgtat
     4261 aagggagcct gacatttata ttccccagaa catcaggtta atggcgtttt tgatgtcatt
     4321 ttcgcggtgg ctgagatcag ccacttcttc cccgataacg gagaccggca cactggccat
     4381 atcggtggtc atcatgcgcc agctttcatc cccgatatgc accaccgggt aaagttcacg
     4441 ggagacttta tctgacagca gacgtgcact ggccaggggg atcaccatcc gtcgcccggg
     4501 cgtgtcaata atatcactct gtacatccac aaacagacga taacggctct ctcttttata
     4561 ggtgtaaacc ttaaactgca tttcaccagc ccctgttctc gtcagcaaaa gagccgttca
     4621 tttcaataaa ccgggcgacc tcagccatcc cttcctgatt ttccgctttc cagcgttcgg
     4681 cacgcagacg acgggcttca ttctgcatgg ttgtgcttac cagaccggag atattgacat
     4741 catatatgcc ttgagcaact gatagctgtc gctgtcaact gtcactgtaa tacgctgctt
     4801 catagcatac ctctttttga catacttcgg gtatacatat cagtatatat tcttataccg
     4861 caaaaatcag cgcgcaaata cgcatactgt tatctggctt ttagtaagcc ggatccacgc
     4921 ggcgtttacg ccccccctgc cactcatcgc agtactgttg taattcatta agcattctgc
     4981 cgacatggaa gccatcacaa acggcatgat gaacctgaat cgccagcggc atcagcacct
     5041 tgtcgccttg cgtataatat ttgcccatgg tgaaaacggg ggcgaagaag ttgtccatat
     5101 tggccacgtt taaatcaaaa ctggtgaaac tcacccaggg attggctgag acgaaaaaca
     5161 tattctcaat aaacccttta gggaaatagg ccaggttttc accgtaacac gccacatctt
     5221 gcgaatatat gtgtagaaac tgccggaaat cgtcgtggta ttcactccag agcgatgaaa
     5281 acgtttcagt ttgctcatgg aaaacggtgt aacaagggtg aacactatcc catatcacca
     5341 gctcaccgtc tttcattgcc atacggaatt ccggatgagc attcatcagg cgggcaagaa
     5401 tgtgaataaa ggccggataa aacttgtgct tatttttctt tacggtcttt aaaaaggccg
     5461 taatatccag ctgaacggtc tggttatagg tacattgagc aactgactga aatgcctcaa
     5521 aatgttcttt acgatgccat tgggatatat caacggtggt atatccagtg atttttttct
     5581 ccattttagc ttccttagct cctgaaaatc tcgacggatc ctaactcaaa atccacacat
     5641 tatacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgcggcc gccatagtga
     5701 ctggatatgt tgtgttttac agtattatgt agtctgtttt ttatgcaaaa tctaatttaa
     5761 tatattgata tttatatcat tttacgtttc tcgttcaact ttattataca tagttgataa
     5821 ttcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa
//
