LOCUS       Exported                8211 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pBCN45-R4R3
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8211)
  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 8211)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8211
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     CDS             3023..3817
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3077..3096)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3687..3706
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4392..4414
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4452..4470)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4538..4642
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4643..5503
                     /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(4861..4880)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5674..6262
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6163..6182
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6388..6410
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6407..6423
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6407..6423
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     protein_bind    6442..6566
                     /gene="mutant version of attR"
                     /label=attR4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(6607..6912)
                     /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(6709..6728)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(7262..7915)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGG"
     primer_bind     7830..7849
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(7969..7999)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(8024..8147)
                     /gene="mutant version of attR"
                     /label=attR3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(8155..8172)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(8155..8171)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8164..8186)
                     /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 gccccatttt atatctgagt
      181 agtatccttt gctttaaatg tccataaaac taattttata atcaataaaa caacgtttgt
      241 aaatcaactg agtttacaag tagagacatt gagggatact ttcactatgc taaagtgata
      301 atcgaccaaa taataacttc actttggtat ttattcctgt cttataaatg ttatgtatga
      361 attaaattca tatgcatatg gctcactctg acaattttta ataatcttcc agatcaatat
      421 tgactaccga tgcgggtggt cttttgcttt gaattctgct gaactttaca ccccgaacag
      481 caatgtgtgc ttcagcctaa aaaaaagtaa gtgtgttaat cagtgccccc gattcttcat
      541 tttttgcccc tctctcccgt ttcgtcggca aaagaagaga aaataaagat aagtctcaag
      601 ataggttggt aatcgctaaa gtggttgtgt ggataagagt agcaaaatgg caggaagagc
      661 actttgcgcg cacacactgt actcattgtt ctggataaaa ttctctcgtt gtttgccgtc
      721 ggatgtctgc ctctctgcat tgagccggct tcttcactat ctttagttaa cctaaaatgc
      781 cgtttctttt ctcgtatccc cactatcccg ttgaggttct ctgctctctt cgctccctac
      841 cgccagcgag caactatccg tgggggcgcc ttgctcggaa gatggggggg aagaaagaag
      901 atttttgcta tttgcacttg agaaagagac ttttcctgcg tcgatggtta gagaacagtg
      961 tgcagacact tttcagctac ctagaattac aattggatat ccccgcctcc caatccaccc
     1021 acccagggaa aaagaagggc tcgccgaaaa tcaaagttat ctccaggctc gcgcatccca
     1081 ccgagcggtt gacttctctc caccactttt cattttaacc ctcgatcgtc agacacagaa
     1141 atggtctcaa agggtgaaga agataacatg gcaattatta aagagtttat gcgtttcaag
     1201 gtgcatatgg agggatctgt caatgggcat gagtttgaaa ttgaaggtga aggagaaggc
     1261 cgaccatatg agggaacaca aaccgcaaaa ctaaaggtaa gtttaaacat atatatacta
     1321 actaaccctg attatttaaa ttttcaggta actaaaggcg gaccattacc attcgcctgg
     1381 gacatcctct ctccacagtt catgtatgga agtaaagctt atgttaaaca tccggcagat
     1441 ataccagatt atttgaaact ttcattcccg gagggtttta agtgggaacg cgtaatgaat
     1501 tttgaagacg gaggagttgt tacagtgacg caagactcaa ggtaagttta aacagttcgg
     1561 tactaactaa ccatacatat ttaaattttc agcctccaag atggagaatt tatttataaa
     1621 gtcaaacttc gaggaacgaa tttcccctcg gatggacctg ttatgcagaa gaagactatg
     1681 ggatgggaag cttcaagtga aagaatgtac cctgaagacg gtgctcttaa gggagagatt
     1741 aaacaacgtc ttaaattgaa agatggagga cattacgatg ctgaggtaag tttaaacatg
     1801 attttactaa ctaactaatc tgatttaaat tttcaggtga agacaactta caaagccaaa
     1861 aaaccagttc agctgccagg agcgtacaat gttaatatta aactggatat cacctcccac
     1921 aacgaggatt acactatcgt tgagcaatat gaaagagctg aagggcggca ctcgacaggt
     1981 ggcatggatg aattgtataa gtagatcaac tgctctaatt ccatctcgtt tctctctttt
     2041 ctttcaatct ctttacccgt ttctcttccc ctacgtcccc caaacctacc actgcacagt
     2101 aacacataca cattcaacgt gtcacaataa aagcccatca acaccttctc tgttgttttt
     2161 tttcttcaaa tgcatttttt tcactttgtt taataatttt tacgcatcgg atttgtagga
     2221 tagtggaaat ttgtccttca atgcttcaat tgattcgaca cttctcaata tttttgatca
     2281 ataaaccatt gattttctat ttagaagaag atcactgtaa ataacaccaa aaacatcaca
     2341 gcaaaaaaac tcccactcaa ttatattact gtttttattg aaactagaat aaatttgaag
     2401 ggcccccctc gaggggagaa aatgcggaga gaagaaccca gcatcttaat aaaattatat
     2461 aattttgaat aaaatcaaat taaaaataag attaattgaa acacaacaat agaatgcaga
     2521 aaattactca taagaaataa tttattttga aagaagttgt gtcgatttac gataaaaact
     2581 acacgcaaac gaggctccgc gtttacaccg tttggcaatg gagcgcgctt gctattagat
     2641 tttaggccag ctcaacgaaa gtttaattta tttttcttct ttttttccat ttttttgagt
     2701 aaaatatcga attttttgta ttaattctgt aaattttata atttgcaact attttggcgt
     2761 attctgcttg tttttatcag cgaattttta tgcatttcag tcaaaagttt gagcaggaaa
     2821 aatgacattt aggactttgt aaaattgtat tcttaaattt gcttgaaaac ctacgtgtac
     2881 aattgatttt attgaatttt tcttacgttt cccgttgtat cttattgtat ttaactttca
     2941 tcataaaata atttatttcc aggtaaaata tccgacgctc tcgtggatcc ccgggattgg
     3001 ctcagaaaaa aaggtaccga ccatgattga acaagatgga ttgcacgcag gttctccggc
     3061 cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga
     3121 tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct
     3181 gtccggtgcc ctgaatgaac tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac
     3241 gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct
     3301 attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt
     3361 atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt
     3421 cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt
     3481 cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag
     3541 gctcaaggcg agcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt
     3601 gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg
     3661 tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg
     3721 cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg
     3781 catcgccttc tatcgccttc ttgacgagtt cttctgagct agccggccat acaagtaatc
     3841 cggatgatcg acgccaacgt cgttgaattt tcaaatttta aatactgaat atttgttttt
     3901 tttcctatta tttatttatt ctctttgtgt tttttttctt gctttctaaa aaattaattc
     3961 aatccaaatc taaacatttt tttttctctt tccgtctccc aattcgtatt ccgctcctct
     4021 catctgaaca caatgtgcaa gtttatttat cttctcgctt tcatttcatt aggacgtggg
     4081 gggaattggt ggaaggggga aacacacaaa aggatgatgg aaatgaaata aggacacaca
     4141 atatgcaaca acattcaatt cagaaatatg gaggaaggtt taaaagaaaa cataaaaata
     4201 tatagaggag gaaggaaaac tagtccaaga tctcagtggt cctaggtgag ctcccatatg
     4261 gtgcactctc agtgcaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc
     4321 aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc
     4381 tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc
     4441 gagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt
     4501 ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     4561 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
     4621 ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
     4681 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
     4741 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
     4801 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
     4861 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
     4921 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
     4981 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
     5041 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
     5101 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     5161 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
     5221 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
     5281 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
     5341 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
     5401 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     5461 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
     5521 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     5581 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     5641 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     5701 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     5761 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     5821 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     5881 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     5941 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     6001 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     6061 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     6121 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     6181 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     6241 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggtctt
     6301 ttgctggcct ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     6361 attaccgcct ttgagtgagc tgataccagc ggataacaat ttcacacagg aaacagctat
     6421 gaccatgatt acgccaagct atcaactttg tatagaaaag ttgaacgaga aacgtaaaat
     6481 gatataaata tcaatatatt aaattagatt ttgcataaaa aacagactac ataatactgt
     6541 aaaacacaac atatccagtc actatggtcg acctgcagac tggctgtgta taagggagcc
     6601 tgacatttat attccccaga acatcaggtt aatggcgttt ttgatgtcat tttcgcggtg
     6661 gctgagatca gccacttctt ccccgataac ggagaccggc acactggcca tatcggtggt
     6721 catcatgcgc cagctttcat ccccgatatg caccaccggg taaagttcac gggagacttt
     6781 atctgacagc agacgtgcac tggccagggg gatcaccatc cgtcgcccgg gcgtgtcaat
     6841 aatatcactc tgtacatcca caaacagacg ataacggctc tctcttttat aggtgtaaac
     6901 cttaaactgc atttcaccag cccctgttct cgtcagcaaa agagccgttc atttcaataa
     6961 accgggcgac ctcagccatc ccttcctgat tttccgcttt ccagcgttcg gcacgcagac
     7021 gacgggcttc attctgcatg gttgtgctta ccagaccgga gatattgaca tcatatatgc
     7081 cttgagcaac tgatagctgt cgctgtcaac tgtcactgta atacgctgct tcatagcata
     7141 cctctttttg acatacttcg ggtatacata tcagtatata ttcttatacc gcaaaaatca
     7201 gcgcgcaaat acgcatactg ttatctggct tttagtaagc cggatccacg cggcgtttac
     7261 gccccccctg ccactcatcg cagtactgtt gtaattcatt aagcattctg ccgacatgga
     7321 agccatcaca aacggcatga tgaacctgaa tcgccagcgg catcagcacc ttgtcgcctt
     7381 gcgtataata tttgcccatg gtgaaaacgg gggcgaagaa gttgtccata ttggccacgt
     7441 ttaaatcaaa actggtgaaa ctcacccagg gattggctga gacgaaaaac atattctcaa
     7501 taaacccttt agggaaatag gccaggtttt caccgtaaca cgccacatct tgcgaatata
     7561 tgtgtagaaa ctgccggaaa tcgtcgtggt attcactcca gagcgatgaa aacgtttcag
     7621 tttgctcatg gaaaacggtg taacaagggt gaacactatc ccatatcacc agctcaccgt
     7681 ctttcattgc catacggaat tccggatgag cattcatcag gcgggcaaga atgtgaataa
     7741 aggccggata aaacttgtgc ttatttttct ttacggtctt taaaaaggcc gtaatatcca
     7801 gctgaacggt ctggttatag gtacattgag caactgactg aaatgcctca aaatgttctt
     7861 tacgatgcca ttgggatata tcaacggtgg tatatccagt gatttttttc tccattttag
     7921 cttccttagc tcctgaaaat ctcgacggat cctaactcaa aatccacaca ttatacgagc
     7981 cggaagcata aagtgtaaag cctggggtgc ctaatgcggc cgccatagtg actggatatg
     8041 ttgtgtttta cagtattatg tagtctgttt tttatgcaaa atctaattta atatattgat
     8101 atttatatca ttttacgttt ctcgttcaac tttattatac atagttgata attcactggc
     8161 cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta a
//
