LOCUS       sequence-94075-e        5261 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PCR template for amplifying the attL1-em7-kanr-attL2-pA cassette.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5261)
  AUTHORS   Zhao L, Ng ET, Koopman P
  TITLE     A piggyBac transposon- and gateway-enhanced system for efficient BAC
            transgenesis.
  JOURNAL   Dev Dyn. 2014 Jun 12. doi: 10.1002/dvdy.24153.
  PUBMED    24924516
REFERENCE   2  (bases 1 to 5261)
  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: "Dev Dyn. 
            2014 Jun 12. doi: 10.1002/dvdy.24153."
COMMENT     Sequence Label: pBSWt1-Dest
FEATURES             Location/Qualifiers
     source          1..5261
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      4..459
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(91..110)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     301..322
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1203..1327
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        1342..1389
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             1421..2215
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             1421..2215
                     /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(1475..1494)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2085..2104
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     protein_bind    complement(2261..2385)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2435..2454)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2489..2508
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             2655..2672
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3073..3093)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3073..3091)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3112..3128)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3112..3128)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3125..3147)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3136..3152
                     /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(3160..3166,3167..3184,3185..3190))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3205..3226
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3343..3360)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3514..4102)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3594..3613)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4273..5064,5065..5133))
                     /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     4896..4915
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5134..5238)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctgacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
       61 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
      121 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
      181 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
      241 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
      301 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
      361 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
      421 ttaacgcgaa ttttaacaaa atattaacgc ttacaatttc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tggagctcca
      661 ccgcggtggc ggccgctcta gctaggctag ggctgctgtg tgaatggagc ggccgagcat
      721 cctggctcct cctccttccc tgctgccggc ccctcttatt tgagctttgg gaagctgggg
      781 gcagccaggc agctggggta aggagttcaa ggcagcgccc acacccgggg ctctccgcaa
      841 cccgaccgcc tgcctgctcc ccctttcctt ttcccgccct ccctcccacc cactcattca
      901 cccacccacc cagagagagg acggcagccc aggaacccgg gcccgccgcc tcctcgccgc
      961 gatcctggac ttcctcctgt cgcaggagcc ggcttccacg tgtgtcccgg agccggcgtc
     1021 tcagcacacg ctccgccggg agcccgggtg cgtccagcag ccggagcaac ctggggaccg
     1081 aggcccccgg agcgcctggg ccaagtccag cgccgagaat ccgcaggatc gcaggagcgg
     1141 agaaccgtcc gcatccgagc cgcacctcag atcccccggg ctgcagacag caagcggcca
     1201 tcacaagttt gtacaaaaaa gctgaacgag aaacgtaaaa tgatataaat atcaatatat
     1261 taaattagat tttgcataaa aaacagacta cataatactg taaaacacaa catatccagt
     1321 cactatggcg gccgcgaatt cgttgacaat taatcatcgg catagtatat cggcatagta
     1381 taatacgaca aggtgaggaa ctaaacccag gaggcagatc atgattgaac aagatggatt
     1441 gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact gggcacaaca
     1501 gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc gcccggttct
     1561 ttttgtcaag accgacctgt ccggtgccct gaatgaactg caagacgagg cagcgcggct
     1621 atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg tcactgaagc
     1681 gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt catctcacct
     1741 tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc atacgcttga
     1801 tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag cacgtactcg
     1861 gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg ggctcgcgcc
     1921 agccgaactg ttcgccaggc tcaaggcgag catgcccgac ggcgaggatc tcgtcgtgac
     1981 ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt ctggattcat
     2041 cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg ctacccgtga
     2101 tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt acggtatcgc
     2161 cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct tctgagcggg
     2221 actctggggt tcgaaatgac cgacaagctt gcaggtcgac catagtgact ggatatgttg
     2281 tgttttacag tattatgtag tctgtttttt atgcaaaatc taatttaata tattgatatt
     2341 tatatcattt tacgtttctc gttcagcttt cttgtacaaa gtggtgatgg ccgctgactc
     2401 gagtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     2461 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttaa
     2521 ttaattaacg ccgcttctcc ccttgtctgt tccttcctga tgccagttct tactcagcag
     2581 ccttccaagg gagtctgaaa gtagcggaca gggaggagtg ggctctgtca aatgcgagtg
     2641 acaacgcgac aacgcaccac caccaccacc acttacacat tactgagcgc aaccccagag
     2701 tccgatagtt cgtaggacgc gggaggacat ctggggcccc ggggtgtcct agagatttcg
     2761 gaactccacg ggaggaggaa gaggaggttg gctgtaacgt gtgactctac accgggggaa
     2821 agtctgatgc ataaggcctc accaggcgtt gactcttaaa acgtctaggg tagaccaaga
     2881 tgaggactcc gagtggtgct gtggtttagg gacatccccc accccccgcc cccgtgctgc
     2941 ctctgacaag cccgaagagc cgacccgaag aacttccagc tctcggtggg ccctggctca
     3001 ttccaggccc accagaagca gtagggagct ttggaatgag ggattaacac tttgggtacc
     3061 cagcttttgt tccctttagt gagggttaat tgcgcgcttg gcgtaatcat ggtcatagct
     3121 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     3181 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     3241 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     3301 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     3361 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     3421 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     3481 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     3541 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     3601 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     3661 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     3721 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     3781 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     3841 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     3901 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt
     3961 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     4021 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     4081 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     4141 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     4201 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     4261 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     4321 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     4381 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     4441 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     4501 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     4561 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     4621 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     4681 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     4741 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     4801 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     4861 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     4921 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     4981 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     5041 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     5101 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     5161 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     5221 acaaataggg gttccgcgca catttccccg aaaagtgcca c
//
