LOCUS       sequence-82107-e        5117 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 5117)
  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 5117)
  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: pBT296_pBS-U-attP-FRT5-pSV40-Neo-pA-FRT5
FEATURES             Location/Qualifiers
     source          1..5117
                     /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    925..972
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     997..1016
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(1204..1998)
                     /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(1204..1998)
                     /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(1315..1334)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1925..1944
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(2033..2390)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2047..2182
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        2199..2390
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     2347..2367
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        complement(2392..2496)
                     /gene="bla"
                     /label=AmpR promoter
     protein_bind    2666..2765
                     /label=phage phi-C31 attP
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attachment site of phage phi-C31"
     promoter        complement(2932..2950)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2971..2987)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2971..2987)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2984..3006)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2995..3011
                     /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(3019..3025,3026..3043,3044..3049))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3064..3085
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3202..3219)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3373..3961)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3453..3472)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4132..4923,4924..4992))
                     /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     4755..4774
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4993..5097)
                     /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 ctctagagat gatcctgagt aacctgaggc tatggcaggg cctgccgccc
      781 cgacgttggc tgcgagccct gggccttcac ccgaacttgg ggggtggggt ggggaaaagg
      841 aagaaacgcg ggcgtattgg ccccaatggg gtctcggtgg ggtatcgaca gagtgccagc
      901 cctgggaccg aaccccgcgt ttatgaacaa acgacccaac accgtgcgtt ttattctgtc
      961 tttttattgc cgtcatagcg cgggttcctt ccggtattgt ctccttccgt gtttcagtta
     1021 gcctccccct agggtgggcg aagaactcca gcatgagatc cccgcgctgg aggatcatcc
     1081 agccggcgtc ccggaaaacg attccgaagc ccaacctttc atagaaggcg gcggtggaat
     1141 cgaaatctcg tgatggcagg ttgggcgtcg cttggtcggt catttcgaac cccagagtcc
     1201 cgctcagaag aactcgtcaa gaaggcgata gaaggcgatg cgctgcgaat cgggagcggc
     1261 gataccgtaa agcacgagga agcggtcagc ccattcgccg ccaagctctt cagcaatatc
     1321 acgggtagcc aacgctatgt cctgatagcg gtccgccaca cccagccggc cacagtcgat
     1381 gaatccagaa aagcggccat tttccaccat gatattcggc aagcaggcat cgccatgggt
     1441 cacgacgaga tcctcgccgt cgggcatgct cgccttgagc ctggcgaaca gttcggctgg
     1501 cgcgagcccc tgatgctctt cgtccagatc atcctgatcg acaagaccgg cttccatccg
     1561 agtacgtgct cgctcgatgc gatgtttcgc ttggtggtcg aatgggcagg tagccggatc
     1621 aagcgtatgc agccgccgca ttgcatcagc catgatggat actttctcgg caggagcaag
     1681 gtgagatgac aggagatcct gccccggcac ttcgcccaat agcagccagt cccttcccgc
     1741 ttcagtgaca acgtcgagca cagctgcgca aggaacgccc gtcgtggcca gccacgatag
     1801 ccgcgctgcc tcgtcttgca gttcattcag ggcaccggac aggtcggtct tgacaaaaag
     1861 aaccgggcgc ccctgcgctg acagccggaa cacggcggca tcagagcagc cgattgtctg
     1921 ttgtgcccag tcatagccga atagcctctc cacccaagcg gccggagaac ctgcgtgcaa
     1981 tccatcttgt tcaatcatgc gaaacgatcc tcatcctgtc tcttgatcga tctttgcaaa
     2041 agcctaggcc tccaaaaaag cctcctcact acttctggaa tagctcagag gccgaggcgg
     2101 cctcggcctc tgcataaata aaaaaaatta gtcagccatg gggcggagaa tgggcggaac
     2161 tgggcggagt taggggcggg atgggcggag ttaggggcgg gactatggtt gctgactaat
     2221 tgagatgcat gctttgcata cttctgcctg ctggggagcc tggggacttt ccacacctgg
     2281 ttgctgacta attgagatgc atgctttgca tacttctgcc tgctggggag cctggggact
     2341 ttccacaccc taactgacac acattccaca gctggttctt tccgcctcag gactcttcct
     2401 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     2461 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     2521 tgaataatcg ctagcgaagt tcctatacct tttgaagaat aggaacttcg gaataggaac
     2581 ttcgtcgaca aaggatcctt ctcgactagt cgcgctcgcg cgactgacgg tcgtaagcac
     2641 ccgcgtacgt gtccaccccg gtcacaaccc cttgtgtcat gtcggcgacc ctacgccccc
     2701 aactgagaga actcaaaggt taccccagtt ggggcactac tcccgaaaac cgcttctgac
     2761 ctgggaaaac gtgaagcccc ggggcatccg ctgagggttg ccgccggggc ttcggtgtgt
     2821 ccgtcagtac tagtcgagaa aagatccgat atccttacgg aataccactt gccacctatc
     2881 accagatctg gatgcatgat ctcctattta aatcagctcc agcttttgtt ccctttagtg
     2941 agggttaatt tcgagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta
     3001 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     3061 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     3121 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     3181 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     3241 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     3301 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     3361 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     3421 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     3481 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     3541 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     3601 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     3661 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     3721 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     3781 gaagtggtgg cctaactacg gctacactag aaggacagta tttggtatct gcgctctgct
     3841 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     3901 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     3961 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     4021 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     4081 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     4141 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     4201 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     4261 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     4321 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     4381 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     4441 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     4501 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     4561 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     4621 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     4681 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     4741 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     4801 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     4861 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     4921 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     4981 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     5041 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     5101 atttccccga aaagtgc
//
