LOCUS       sequence-313563-        4568 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 4568)
  AUTHORS   Perez-Pinera P, Ousterout DG, Brown MT, Gersbach CA
  TITLE     Gene targeting to the ROSA26 locus directed by engineered zinc 
            finger nucleases.
  JOURNAL   Nucleic Acids Res. 2012 Apr 1;40(8):3741-52. Epub 2011 Dec 14.
  PUBMED    22169954
REFERENCE   2  (bases 1 to 4568)
  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: "Nucleic 
            Acids Res."; date: "2012-04-1"; volume: "40(8)"; pages: "3741-52. 
            Epub 2011 Dec 14"
COMMENT     Sequence Label: pDonor MCS Rosa26
FEATURES             Location/Qualifiers
     source          1..4568
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    65..858
                     /label=Rosa26 left arm
                     /note="left homology arm from intron 1 of the mouse Rosa26 
                     gene (Soriano, 1999)"
     misc_feature    906..1716
                     /label=Rosa26 right arm
                     /note="right homology arm from intron 1 of the mouse Rosa26
                     gene (Soriano, 1999)"
     primer_bind     complement(1743..1763)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1743..1761)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1782..1798)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1782..1798)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1795..1817)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1806..1822
                     /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(1830..1836,1837..1854,1855..1860))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1875..1896
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2013..2030)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2184..2772)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2264..2283)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2943..3734,3735..3803))
                     /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     3566..3585
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3804..3908)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3934..4389)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4071..4092)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4283..4302
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4516..4538
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4530..4547
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4531..4547
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     join(4557..4568,1..8)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        join(4557..4568,1..7)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ctatagggcg aattgggtac ctgaaaacac aaatggcgtg ttttggttgg cgtaaggcgc
       61 ctgtcagtta acggcagccg gagtgcgcag ccgccggcag cctcgctctg cccactgggt
      121 ggggcgggag gtaggtgggg tgaggcgagc tggacgtgcg ggcgcggtcg gcctctggcg
      181 gggcggggga ggggagggag ggtcagcgaa agtagctcgc gcgcgagcgg ccgcccaccc
      241 tccccttcct ctgggggagt cgttttaccc gccgccggcc gggcctcgtc gtctgattgg
      301 ctctcggggc ccagaaaact ggcccttgcc attggctcgt gttcgtgcaa gttgagtcca
      361 tccgccggcc agcgggggcg gcgaggaggc gctcccaggt tccggccctc ccctcggctc
      421 cgcgccgcag agtctggccg cgcgcccctg cgcaacgtgg caggaagcgc gcgctggggg
      481 cggggacggg cagtagggct gagcggctgc ggggcgggtg caagcacgtt tccgacttga
      541 gttgcctcaa gaggggcgtg ctgagccaga cctccatcgc gcactccggg gagtggaggg
      601 aaggagcgag ggctcagttg ggctgttttg gaggcaggaa gcacttgctc tcccaaagtc
      661 gctctgagtt gttatcagta agggagctgc agtggagtag gcggggagaa ggccgcaccc
      721 ttctccggag gggggagggg agtgttgcaa tacctttctg ggagttctct gctgcctcct
      781 ggcttctgag gaccgccctg ggcctgggag aatcccttcc ccctcttccc tcgtgatctg
      841 caactccagt ctttctagag aattctctag aaatattctc gaggtttaaa cgtcgacgcg
      901 gccgcagatg ggcgggagtc ttctgggcag gcttaaaggc taacctggtg tgtgggcgtt
      961 gtcctgcagg ggaattgaac aggtgtaaaa ttggagggac aagacttccc acagattttc
     1021 ggttttgtcg ggaagttttt taataggggc aaataaggaa aatgggagga taggtagtca
     1081 tctggggttt tatgcagcaa aactacaggt tattattgct tgtgatccgc ctcggagtat
     1141 tttccatcga ggtagattaa agacatgctc acccgagttt tatactctcc tgcttgagat
     1201 ccttactaca gtatgaaatt acagtgtcgc gagttagact atgtaagcag aattttaatc
     1261 atttttaaag agcccagtac ttcatatcca tttctcccgc tccttctgca gccttatcaa
     1321 aaggtatttt agaacactca ttttagcccc attttcattt attatactgg cttatccaac
     1381 ccctagacag agcattggca ttttcccttt cctgatctta gaagtctgat gactcatgaa
     1441 accagacaga ttagttacat acaccacaaa tcgaggctgt agctggggcc tcaacactgc
     1501 agttctttta taactcctta gtacactttt tgttgatcct ttgccttgat ccttaatttt
     1561 cagtgtctat cacctctccc gtcaggtggt gttccacatt tgggcctatt ctcagtccag
     1621 ggagttttac aacaatagat gtattgagaa tccaacctaa agcttaactt tccactccca
     1681 tgaatgcctc tctccttttt ctccatttat aaactgccgc ggtggagctc cagcttttgt
     1741 tccctttagt gagggttaat tgcgcgcttg gcgtaatcat ggtcatagct gtttcctgtg
     1801 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa
     1861 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct
     1921 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     1981 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     2041 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     2101 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     2161 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     2221 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     2281 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     2341 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     2401 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     2461 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     2521 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     2581 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc
     2641 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     2701 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     2761 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     2821 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     2881 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     2941 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     3001 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     3061 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     3121 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     3181 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     3241 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     3301 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     3361 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     3421 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     3481 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     3541 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     3601 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     3661 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     3721 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     3781 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     3841 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     3901 gttccgcgca catttccccg aaaagtgcca cctaaattgt aagcgttaat attttgttaa
     3961 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
     4021 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
     4081 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
     4141 agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc
     4201 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
     4261 cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg
     4321 caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac
     4381 agggcgcgtc ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg
     4441 cctcttcgct attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg
     4501 taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggccagtgag cgcgcgtaat
     4561 acgactca
//
