LOCUS       sequence-12435-e        7999 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 7999)
  AUTHORS   Giordano-Santini R, Milstein S, Svrzikapa N, Tu D, Johnsen R, 
            Baillie D, Vidal M, Dupuy D
  TITLE     An antibiotic selection marker for nematode transgenesis.
  JOURNAL   Nat Methods. 2010 Sep;7(9):721-3. doi: 10.1038/nmeth.1494. Epub 2010
            Aug 22.
  PUBMED    20729841
REFERENCE   2  (bases 1 to 7999)
  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.1494"; journalName: "Nat Methods"; date: "2010-09"; 
            volume: "7"; issue: "9"; pages: "721-3"
COMMENT     Sequence Label: pRG5271Neo
FEATURES             Location/Qualifiers
     source          1..7999
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             795..1589
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(849..868)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1459..1478
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     2290..2306
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     2291..2307
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        complement(3686..3704)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3725..3741)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3725..3741)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3738..3760)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3749..3765
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(3773..3779,3780..3797,3798..3803))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3818..3839
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3956..3973)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4127..4715)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4207..4226)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4886..5677,5678..5746))
                     /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     5509..5528
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5747..5851)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5877..6332)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6014..6035)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6226..6245
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6459..6481
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     6473..6490
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6474..6490
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6497..6516
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        6497..6515
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 atttctaaat tttagtttaa taaaaaaatt ttaaaataat acgagttttg agactaactt
       61 tcaataaaaa ccaaaaaatt gcttcaaaaa agagctcgcg ccgatttacg ataaacacag
      121 cgcgctacga cactccgaaa accgcgccgc accattacat tctccgcccc accgcacggt
      181 ttgcggaagg cgatgaaatt gctgggccta aaattttaac cctttttttc ccgtttttca
      241 actgttttcc cgattttttc gatatttttt tcaggttttt agttagaaaa gtatataaaa
      301 attgcattat ttgtagtttt cgctcgtaat ttgtgataga aaacggccaa tttgcctaga
      361 tttgtgattc taccaagtgg aatttttggg aaaatatcaa tattccacga tttttattca
      421 aatcataacg tttcgaaatt atcagcttaa ataaaactaa aataaataat aaaatactaa
      481 aaaccctgaa tttctcggtt ctcggttcag cttgagttcg ttctatacac gttatcgatt
      541 tttgtcattt cgtttaatca tgggttttga ttcattttaa atatgaaatt tgatttattc
      601 ctcgtttcga agttggtttg tcttgtccgg ccggtaaaca aatgttatta aaaaaaataa
      661 taaatttcga aatttttcgt aatgttccgc aatcgtgttt ttccgtttat ctcgtcttcc
      721 cttccttcaa atatactatt tattttctca attcacatat tttcagccag ccctgctcaa
      781 caagtggaat aaaaatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg
      841 tggagaggct attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg
      901 tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg
      961 ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc
     1021 cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg
     1081 aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca
     1141 tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc
     1201 aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg
     1261 atgatctgga cgaagagcat caggggctcg caccagccga actgttcgcc aggctcaagg
     1321 cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata
     1381 tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg
     1441 accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat
     1501 gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct
     1561 tctatcgcct tcttgacgag ttcttctgag tccaattact cttcaacatc cctacatgct
     1621 ctttctccct gtgctcccac cccctatttt tgttattatc aaaaaacttc tcttaatttc
     1681 tttgtttttt agcttctttt aagccacctc taacaatgaa attgtgtaga ttcaaaaata
     1741 gaattaattc gtaataaaaa gtcgaaaaaa attgtgctcc ctccccccat taataataat
     1801 tctatcccaa aatctacaca atgttctgtg tacacttctt atgtttttta cttctgataa
     1861 atttttttga aacatcatag aaaaaaccgc acacaaaata ccttatcata tgttacgttt
     1921 cagtttatga ccgcaatttt tatttcttcg cacgtctggg cctctcatga cgtcaaatca
     1981 tgctcatcgt gaaaaagttt tggagtattt ttggaatttt tcaatcaagt gaaagtttat
     2041 gaaattaatt ttcctgcttt tgctttttgg ggtttcccct attgtttgtc aagatttcga
     2101 ggacggcgtt tttcttgcta aaatcacaag tattgatgag cacgatgcaa gaaagatcgg
     2161 aagaaggttt gggtttgagg ctcagtggaa ggtgagtaga agttgataat ttgaaagtgg
     2221 agtagtgtct atggggtttt tgccttaaat gacagaatac attcccaata taccaaacat
     2281 aactgtttct cgaggtcgac ggtatcgata agcttgatat cgaattcctg cagcccgggt
     2341 atatatatat tgggatgcac tctcaatggt cctgtaacac ttttttgtgt gttagtgacc
     2401 agtgctccgc tcccagtaga tgtttttgca caaaaacata ccaaaatgtt ttaccatatt
     2461 catagatcat tgtcatagaa atttgtttaa ttctgaaaag tttcctctgt tttatggcag
     2521 aaacaattca ttgagataat ctgacttatt attcattggg ggtaagtgct ctatgggatc
     2581 cctttaatcg acgattaacc ttcaaacctc aaaaaaaaaa aacaagaaaa tcacaaagct
     2641 tttcctatct tcacaaagct tttcctatct taaaggcata actgtgtgtg tatatctgat
     2701 ctttgtccaa ctgtatagat gtgttttgtt tctgagaaca aaagttgtcc atggaccaaa
     2761 tttctataga agttgcacgc actgtacttc tagattttag gtagagtgtg tgattgaaag
     2821 ttatattcag caaaattata atcagtgatc ctttccaaag cttggcattg ttgcttgaaa
     2881 tcctgttcaa gtagtgtttg gcaaaaaatt cccaaaattt ccgatatgta ctctatgttt
     2941 aattttgggt tttaaataaa tacgttgaat tacgcaagct gttacttcta aaaaacattg
     3001 tatacatttt agggttgtag tgaactttta taatatttgc tcaaaaactc tgaaacatta
     3061 aaatgacgtg ctggtctgct tttattctcg aattatatga tcaaatctct gattttctgg
     3121 ttaaaaaatt ccctattgta ttttgataga ctttttctcc ttatttcatt ttttcacagg
     3181 gtatatgaga tgatgttcaa agtggttcat aatatccagt taagaatcaa aatacttcac
     3241 tttcaaatta agtgtcctgg gcttctgaat gcaggtgttg tctacatatt atttctcaaa
     3301 tcaaaatact ctgtcatttc ttacgagctt aaccaattca ccttttgtat ggcagcagca
     3361 ggcgtcatac atttccatct caattgacct attctcgagt ctcggctgtc catgtaatta
     3421 tccattttca ccgtgaaccc ttctgtccat cgtatttttt tcccatcgct cctctctttc
     3481 tcttcatctc cacagtttga cctttacaca gaaactcttt ttcccgaata atatgtagtt
     3541 cttctcgtag ttttcggaag aagtaccgcc cgccgcaagg atttctcttc gttccgtcca
     3601 tcttaaaaac acgagcttaa ttgagaagcg atcatgtctt cttgtcccgg cggccgccac
     3661 cgcggtggag ctccagcttt tgttcccttt agtgagggtt aatttcgagc ttggcgtaat
     3721 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac
     3781 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa
     3841 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat
     3901 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc
     3961 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     4021 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     4081 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     4141 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     4201 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     4261 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     4321 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     4381 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     4441 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     4501 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     4561 ctagaaggac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     4621 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     4681 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     4741 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     4801 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     4861 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     4921 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     4981 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     5041 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     5101 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     5161 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     5221 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     5281 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     5341 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     5401 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     5461 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     5521 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     5581 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     5641 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     5701 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     5761 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     5821 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctaaat
     5881 tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt
     5941 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
     6001 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt
     6061 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc
     6121 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
     6181 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
     6241 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
     6301 cgccgcgctt aatgcgccgc tacagggcgc gtcccattcg ccattcaggc tgcgcaactg
     6361 ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga aagggggatg
     6421 tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac gttgtaaaac
     6481 gacggccagt gaattgtaat acgactcact atagggcgaa ttgggtacca attttcatac
     6541 cttaaactcc tcaaagaaac caataattat ttgaacaaaa cacacaaaaa agtaccaggt
     6601 ttttcaaaac tttcaatcga atattttggt taatttccaa atttcggaac aatttgaagc
     6661 attgctttgt gctctcctat gatattattc ttcacttatc ttgcatggca ttttcactgt
     6721 aatccctata tgtcactttc acatccatca gtttttttaa acattgaatt aatatgtttc
     6781 tagtacttaa ctacatgaac tagttgacct actctctgtg gaaaaaagga caacagcaag
     6841 tagaatggga ggaagtttct ctttttctta ttttcattga aaagggtgaa aacgaggttg
     6901 ttgctccatt tccgactgtc gtcaagactg tacctacagg ataattacgc aatgttgttc
     6961 tcccgcacag caaaaaaaaa gagaaaagtc gaatggtttg tgtgacacct tgagatgccg
     7021 catgatcgat gaaccttcac tcaaataaac cgaaaagtaa attatcgaaa tgaaaggagc
     7081 gagcaaaaaa aagaaataga gggcagttca acgaaaactt tccacttctt ttgatctatt
     7141 atattgcagg tatcaaaatc aggatttaag ggaaatgtgt tctgtgcttc cgatatactg
     7201 agcgttcatg gattaagaag acaatattga gaaggaatca gctagatttt ccgatatcaa
     7261 aatttcagac caaacaaaaa ttgaaaaaaa aatcgtaatt ccattaatat acttcttgta
     7321 gatcttatat ataggtagga ataaagacca acttatgtgc ctttaagcag gcgtaactcg
     7381 ctttcttggc gtttgggggc aacttactgc acaacatcaa ttactttcta ccttcagcct
     7441 gacaacttat taaaagctga tcaaataaat tggtagtcct ggcaagaaac ggccaaccaa
     7501 aaatgtttaa cacgaatttt tcaaatttgg caaaagcttt cactcaactt ctaagcttac
     7561 atcaccgtat agaaagatat cacacggact ggttaggtct ttttcaagcc gtttaggcga
     7621 gtaggcaggt attttgtgcc tacagtgcgt gcaacttcta tagaaccccc taaattttcg
     7681 agtttcagca tgttctgata tttttcaaga aaatttgtta atgtcaatcg attctaaatg
     7741 ccaaaaaacc tactgtccaa aatttttaga aaaatatatg caaaactacg agaattaggc
     7801 caaaaagtcg taaaaacgac cgtgcaactt ctatagaacc cctcagaatt ttgattttaa
     7861 tcgcaaaaag cactgaattt gtttaaattt ttcaaaataa ttttgtaaaa caataacatc
     7921 aaaatgatct aaatacttag ttcgttcatt aaaaacatga aaaatgctac gtggaaatat
     7981 atatatatat atagggccc
//
