LOCUS       sequence-82319-e        7207 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of FRT5-flanked Neo; homologous recombination targeting 
            vector cloning.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7207)
  AUTHORS   Tasic B, Hippenmeyer S, Wang C, Gamboa M, Zong H, Chen-Tsai Y, Luo L
  TITLE     Site-specific integrase-mediated transgenesis in mice via pronuclear
            injection.
  JOURNAL   Proc Natl Acad Sci U S A. 2011 May 10;108(19):7902-7. doi: 
            10.1073/pnas.1019507108. Epub 2011 Apr 4.
  PUBMED    21464299
REFERENCE   2  (bases 1 to 7207)
  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: 
            "10.1073/pnas.1019507108"; journalName: "Proc Natl Acad Sci U S A"; 
            date: "2011-05-10- 10"; volume: "108"; issue: "19"; pages: "7902-7"
COMMENT     Sequence Label: 
            pBT307_pBS-U-attPx3-FRT5-pSV40-Neo-pA-PhiC31o-pA-FRT5
FEATURES             Location/Qualifiers
     source          1..7207
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(33..461)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(143..164)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     355..374
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     588..610
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     602..619
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     603..619
                     /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"
     polyA_signal    742..966
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     955..972
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(999..1019)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(1020..2834)
                     /codon_start=1
                     /product="integrase from phage phi-C31"
                     /label=phage phi-C31 integrase
                     /translation="MDTYAGAYDRQSRERENSSAASPATQRSANEDKAADLQREVERDG
                     GRFRFVGHFSEAPGTSAFGTAERPEFERILNECRAGRLNMIIVYDVSRFSRLKVMDAIP
                     IVSELLALGVTIVSTQEGVFRQGNVMDLIHLIMRLDASHKESSLKSAKILDTKNLQREL
                     GGYVGGKAPYGFELVSETKEITRNGRMVNVVINKLAHSTTPLTGPFEFEPDVIRWWWRE
                     IKTHKHLPFKPGSQAAIHPGSITGLCKRMDADAVPTRGETIGKKTASSAWDPATVMRIL
                     RDPRIAGFAAEVIYKKKPDGTPTTKIEGYRIQRDPITLRPVELDCGPIIEPAEWYELQA
                     WLDGRGRGKGLSRGQAILSAMDKLYCECGAVMTSKRGEESIKDSYRCRRRKVVDPSAPG
                     QHEGTCNVSMAALDKFVAERIFNKIRHAEGDEETLALLWEAARRFGKLTEAPEKSGERA
                     NLVAERADALNALEELYEDRAAGAYDGPVGRKHFRKQQAALTLRQQGAEERLAELEAAE
                     APKLPLDQWFPEDADADPTGPKSWWGRASVDDKRVFVGLFVDKIVVTKSTTGRGQGTPI
                     EKRASITWAKPPTDDDEDDAQDGTEDVAA"
     regulatory      2831..2840
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     polyA_signal    3054..3101
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     3126..3145
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(3333..4127)
                     /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             complement(3333..4127)
                     /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(3444..3463)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4054..4073
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(4162..4519)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4176..4311
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        4328..4519
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     4476..4496
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        complement(4521..4625)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        complement(5022..5040)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5061..5077)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5061..5077)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5074..5096)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5085..5101
                     /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(5109..5115,5116..5133,5134..5139))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5154..5175
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5292..5309)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5463..6051)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5543..5562)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6222..7013,7014..7082))
                     /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     6845..6864
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7083..7187)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
       61 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
      121 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
      181 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
      241 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
      301 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
      361 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
      421 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc cattcaggct
      481 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
      541 agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg
      601 ttgtaaaacg acggccagtg aattgtaata cgactcacta tagggcgaat tgggtacagg
      661 cgcgcccctt actaggcggc cgcgaagttc ctataccttt tgaagaatag gaacttcgga
      721 ataggaactt ctctaggacg tccatagagc ccaccgcatc cccagcatgc ctgctattgt
      781 cttcccaatc ctcccccttg ctgtcctgcc ccaccccacc ccccagaata gaatgacacc
      841 tactcagaca atgcgatgca atttcctcat tttattagga aaggacagtg ggagtggcac
      901 cttccagggt caaggaaggc acgggggagg ggcaaacaac agatggctgg caactagaag
      961 gcacagtcga ggctgatcag cgagctctag atctatcaca ctttccgctt tttcttaggg
     1021 gcggccacgt cctcggtgcc gtcctgggcg tcatcctcgt cgtcgtcggt gggaggcttg
     1081 gcccaggtga tgctggctct cttctcgatg ggggtgccct ggcccctgcc ggtggtgctc
     1141 ttggtcacca cgatcttatc cacgaacagg cccacgaaca ccctcttgtc gtccacgctg
     1201 gctctgcccc accaggactt ggggccggtg gggtcggcgt cggcatcctc ggggaaccac
     1261 tgatccaggg gcagcttagg ggcctcggcg gcctccagct cggccagtct ttcctcggcg
     1321 ccctgctgtc tcagggtcag ggcggcctgc tgctttctga agtgcttcct gcccacaggg
     1381 ccgtcatagg ctccggcggc tctgtcctcg tacagctcct ccagggcgtt cagggcgtcg
     1441 gctctctcgg ccaccaggtt ggccctctcg ccgctcttct cgggggcctc ggtcagcttg
     1501 ccgaatcttc tggcggcctc ccacagcagg gccagggtct cctcgtcgcc ctcggcgtgc
     1561 cggatcttgt tgaagatccg ctcggccacg aacttgtcca gggcggccat gctcacatta
     1621 caggtgccct cgtgctggcc aggggcgctg gggtccacca cctttcttct ccggcaccgg
     1681 tagctgtcct tgatgctctc ctcgcctctc ttgctggtca tcacggcgcc acactcacag
     1741 tacagcttgt ccatggcgct caggatggcc tggcctctgc tcaggccctt gcctctgcct
     1801 ctgccgtcca gccaggcctg cagctcgtac cactcggcag gctcgatgat agggccacag
     1861 tccagctcca caggtctcag ggtgatgggg tctctctgga ttctgtagcc ctcgatcttg
     1921 gtggtggggg tgccgtcggg cttcttcttg tagatcacct cggcggcgaa gccggcgatc
     1981 ctagggtccc tcaggattct catcacggtg gcggggtccc aggcgctgct ggcggttttc
     2041 ttgccgatgg tctcgcctct ggtgggcacg gcgtcggcgt ccattctctt acacaggccg
     2101 gtgatgctgc cggggtggat ggcggcctgg ctgccgggct tgaaaggcag gtgcttgtgg
     2161 gtcttgatct cccgccacca ccacctgatc acgtcgggct caaactcgaa ggggccggtc
     2221 aggggggtgg tgctgtgggc cagcttgttg atcaccacgt tcaccatcct gccgttccgg
     2281 gtgatctcct tggtctcgga caccagctcg aagccgtagg gggccttgcc gcccacatag
     2341 ccgcccagct ccctctgcag gttcttggtg tccaggatct tggcgctctt caggctgctc
     2401 tccttgtggc tggcgtccag cctcatgatc aggtggatca ggtccatcac gttgccctgt
     2461 ctaaagacgc cttcctgggt ggacacgatg gtcacgccca gggccagcag ctcggacacg
     2521 atagggatgg cgtccatcac cttcagcctg ctgaaccggg acacgtcgta cacgatgatc
     2581 atgttcagcc tgccggccct acactcgttc aggattctct cgaactcggg tctctcggcg
     2641 gtgccgaagg cgctggtgcc aggggcctcg ctgaagtggc ccacaaatct gaatctgccg
     2701 ccgtccctct ccacctctct ctgcagatcg gcggccttat cctcgttggc gcttctctgg
     2761 gtggcggggc tggcggcgct gctgttctct ctctcccggc tctgtctgtc gtaggctccg
     2821 gcgtaggtat ccatggtggc ggttaactta attcctgcag gctagagatg atcctgagta
     2881 acctgaggct atggcagggc ctgccgcccc gacgttggct gcgagccctg ggccttcacc
     2941 cgaacttggg gggtggggtg gggaaaagga agaaacgcgg gcgtattggc cccaatgggg
     3001 tctcggtggg gtatcgacag agtgccagcc ctgggaccga accccgcgtt tatgaacaaa
     3061 cgacccaaca ccgtgcgttt tattctgtct ttttattgcc gtcatagcgc gggttccttc
     3121 cggtattgtc tccttccgtg tttcagttag cctcccccta gggtgggcga agaactccag
     3181 catgagatcc ccgcgctgga ggatcatcca gccggcgtcc cggaaaacga ttccgaagcc
     3241 caacctttca tagaaggcgg cggtggaatc gaaatctcgt gatggcaggt tgggcgtcgc
     3301 ttggtcggtc atttcgaacc ccagagtccc gctcagaaga actcgtcaag aaggcgatag
     3361 aaggcgatgc gctgcgaatc gggagcggcg ataccgtaaa gcacgaggaa gcggtcagcc
     3421 cattcgccgc caagctcttc agcaatatca cgggtagcca acgctatgtc ctgatagcgg
     3481 tccgccacac ccagccggcc acagtcgatg aatccagaaa agcggccatt ttccaccatg
     3541 atattcggca agcaggcatc gccatgggtc acgacgagat cctcgccgtc gggcatgctc
     3601 gccttgagcc tggcgaacag ttcggctggc gcgagcccct gatgctcttc gtccagatca
     3661 tcctgatcga caagaccggc ttccatccga gtacgtgctc gctcgatgcg atgtttcgct
     3721 tggtggtcga atgggcaggt agccggatca agcgtatgca gccgccgcat tgcatcagcc
     3781 atgatggata ctttctcggc aggagcaagg tgagatgaca ggagatcctg ccccggcact
     3841 tcgcccaata gcagccagtc ccttcccgct tcagtgacaa cgtcgagcac agctgcgcaa
     3901 ggaacgcccg tcgtggccag ccacgatagc cgcgctgcct cgtcttgcag ttcattcagg
     3961 gcaccggaca ggtcggtctt gacaaaaaga accgggcgcc cctgcgctga cagccggaac
     4021 acggcggcat cagagcagcc gattgtctgt tgtgcccagt catagccgaa tagcctctcc
     4081 acccaagcgg ccggagaacc tgcgtgcaat ccatcttgtt caatcatgcg aaacgatcct
     4141 catcctgtct cttgatcgat ctttgcaaaa gcctaggcct ccaaaaaagc ctcctcacta
     4201 cttctggaat agctcagagg ccgaggcggc ctcggcctct gcataaataa aaaaaattag
     4261 tcagccatgg ggcggagaat gggcggaact gggcggagtt aggggcggga tgggcggagt
     4321 taggggcggg actatggttg ctgactaatt gagatgcatg ctttgcatac ttctgcctgc
     4381 tggggagcct ggggactttc cacacctggt tgctgactaa ttgagatgca tgctttgcat
     4441 acttctgcct gctggggagc ctggggactt tccacaccct aactgacaca cattccacag
     4501 ctggttcttt ccgcctcagg actcttcctt tttcaatatt attgaagcat ttatcagggt
     4561 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
     4621 ccgcgcacat ttccccgaaa agtgccacct gaataatcgc tagcgaagtt cctatacctt
     4681 ttgaagaata ggaacttcgg aataggaact tcgtcgacaa aggatccgtc ggcgacccta
     4741 cgcccccaac tgagagaact caaaggttac cccagttggg gcactactcc cgaaaacgtc
     4801 ggcgacccta cgcccccaac tgagagaact caaaggttac cccagttggg gcactactcc
     4861 cgaaaacgtc ggcgacccta cgcccccaac tgagagaact caaaggttac cccagttggg
     4921 gcactactcc cgaaaacgat atccttacgg aataccactt gccacctatc accagatctg
     4981 gatgcatgat ctcctattta aatcagctcc agcttttgtt ccctttagtg agggttaatt
     5041 tcgagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca
     5101 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg
     5161 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg
     5221 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     5281 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     5341 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     5401 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     5461 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     5521 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     5581 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     5641 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     5701 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     5761 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     5821 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     5881 cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct gaagccagtt
     5941 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     6001 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     6061 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     6121 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     6181 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     6241 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     6301 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     6361 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     6421 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     6481 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     6541 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     6601 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     6661 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     6721 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     6781 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     6841 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     6901 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     6961 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     7021 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     7081 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     7141 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     7201 aaagtgc
//
