LOCUS       sequence-103227-        4198 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Donor vector for tdTomato knock-in via landing pad.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4198)
  AUTHORS   Ohtsuka M, Miura H, Mochida K, Hirose M, Hasegawa A, Ogura A, 
            Mizutani R, Kimura M, Isotani A, Ikawa M, Sato M, Gurumurthy CB
  TITLE     One-step generation of multiple transgenic mouse lines using an 
            improved Pronuclear Injection-based Targeted Transgenesis (i-PITT).
  JOURNAL   BMC Genomics. 2015 Apr 9;16:274. doi: 10.1186/s12864-015-1432-5.
  PUBMED    25887549
REFERENCE   2  (bases 1 to 4198)
  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: "BMC 
            Genomics."; date: "2015-04-9"; volume: "16:274. doi"; pages: " 
            10.1186/s12864-015-1432-5"
COMMENT     Sequence Label: pBEZ
FEATURES             Location/Qualifiers
     source          1..4198
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     5..22
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     6..22
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    168..201
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="minimal attB site for the phi-C31 integrase (Groth 
                     et al., 2000)"
     protein_bind    221..254
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="minimal attB site for the phi-C31 integrase (Groth 
                     et al., 2000)"
     protein_bind    274..307
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="minimal attB site for the phi-C31 integrase (Groth 
                     et al., 2000)"
     protein_bind    327..360
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="minimal attB site for the phi-C31 integrase (Groth 
                     et al., 2000)"
     protein_bind    380..413
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="minimal attB site for the phi-C31 integrase (Groth 
                     et al., 2000)"
     protein_bind    433..466
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="minimal attB site for the phi-C31 integrase (Groth 
                     et al., 2000)"
     regulatory      534..543
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             540..1970
                     /codon_start=1
                     /product="tandem dimeric (pseudo-monomeric) derivative of 
                     DsRed (Shaner et al., 2004)"
                     /label=tdTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLGHGTGSTGSGSSGTASSEDNNMAVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPY
                     EGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVM
                     NFEDGGLVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGV
                     LKGEIHQALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYE
                     RSEGRHHLFLYGMDELYK"
     polyA_signal    2097..2218
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2134..2153)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2188..2207
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    complement(2334..2381)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     protein_bind    complement(2382..2415)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     primer_bind     complement(2499..2515)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2499..2515)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     terminator      2563..2594
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     promoter        2595..2697
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             2698..3513
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCLKPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNGDRVFRLAQAQSRMNNGLVGA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2771..2790)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      3537..3564
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     rep_origin      complement(3576..4163)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3656..3675)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cccgtgtaaa acgacggcca gtttatctag tcagcttgat tctagctgat cgtggaccgg
       61 aaggtgagcc agtgagttga ttgcagtcca gttacgctgg agtctgaggc tcgtcctgaa
      121 tgatatgcgg cctcggatcc gcgatcgatc cgctagccgc ggtgcgggtg ccagggcgtg
      181 cccttgggct ccccgggcgc gtactccacc cgcggtgcgg gtgccagggc gtgcccttgg
      241 gctccccggg cgcgtactcc acccgcggtg cgggtgccag ggcgtgccct tgggctcccc
      301 gggcgcgtac tccacccgcg gtgcgggtgc cagggcgtgc ccttgggctc cccgggcgcg
      361 tactccaccc gcggtgcggg tgccagggcg tgcccttggg ctccccgggc gcgtactcca
      421 cccgcggtgc gggtgccagg gcgtgccctt gggctccccg ggcgcgtact ccacctagga
      481 agttcctatt ccgaagttcc tattctatca gaagtatagg aacttcaccg gtcgccacca
      541 tggtgagcaa gggcgaggag gtcatcaaag agttcatgcg cttcaaggtg cgcatggagg
      601 gctccatgaa cggccacgag ttcgagatcg agggcgaggg cgagggccgc ccctacgagg
      661 gcacccagac cgccaagctg aaggtgacca agggcggccc cctgcccttc gcctgggaca
      721 tcctgtcccc ccagttcatg tacggctcca aggcgtacgt gaagcacccc gccgacatcc
      781 ccgattacaa gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg
      841 aggacggcgg tctggtgacc gtgacccagg actcctccct gcaggacggc acgctgatct
      901 acaaggtgaa gatgcgcggc accaacttcc cccccgacgg ccccgtaatg cagaagaaga
      961 ccatgggctg ggaggcctcc accgagcgcc tgtacccccg cgacggcgtg ctgaagggcg
     1021 agatccacca ggccctgaag ctgaaggacg gcggccacta cctggtggag ttcaagacca
     1081 tctacatggc caagaagccc gtgcaactgc ccggctacta ctacgtggac accaagctgg
     1141 acatcacctc ccacaacgag gactacacca tcgtggaaca gtacgagcgc tccgagggcc
     1201 gccaccacct gttcctgggg catggcaccg gcagcaccgg cagcggcagc tccggcaccg
     1261 cctcctccga ggacaacaac atggccgtca tcaaagagtt catgcgcttc aaggtgcgca
     1321 tggagggctc catgaacggc cacgagttcg agatcgaggg cgagggcgag ggccgcccct
     1381 acgagggcac ccagaccgcc aagctgaagg tgaccaaggg cggccccctg cccttcgcct
     1441 gggacatcct gtccccccag ttcatgtacg gctccaaggc gtacgtgaag caccccgccg
     1501 acatccccga ttacaagaag ctgtccttcc ccgagggctt caagtgggag cgcgtgatga
     1561 acttcgagga cggcggtctg gtgaccgtga cccaggactc ctccctgcag gacggcacgc
     1621 tgatctacaa ggtgaagatg cgcggcacca acttcccccc cgacggcccc gtaatgcaga
     1681 agaagaccat gggctgggag gcctccaccg agcgcctgta cccccgcgac ggcgtgctga
     1741 agggcgagat ccaccaggcc ctgaagctga aggacggcgg ccactacctg gtggagttca
     1801 agaccatcta catggccaag aagcccgtgc aactgcccgg ctactactac gtggacacca
     1861 agctggacat cacctcccac aacgaggact acaccatcgt ggaacagtac gagcgctccg
     1921 agggccgcca ccacctgttc ctgtacggca tggacgagct gtacaagtaa agcggccggc
     1981 cgcgactcta gatcataatc agccatacca catttgtaga ggttttactt gctttaaaaa
     2041 acctcccaca cctccccctg aacctgaaac ataaaatgaa tgcaattgtt gttgttaact
     2101 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     2161 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttaag
     2221 gcgtaaattg taagcgttaa tgatccacta gaggatcccc gggtaccgag ctcgaattcg
     2281 aagttcctat actcctataa gaataggaac ttcggaatag gaacttcatg atggaagttc
     2341 atatactttc tagagaatag gaacttcgga ataggaactt cataacttcg tataggatac
     2401 tttatacgaa gttatcagct gccgctcgag gcgcgtgatc ttacggcatt atacgtatga
     2461 tcggtccacg atcagctaga ttatctagtc agcttgatgt catagctgtt tcctgaggct
     2521 caatactgac catttaaatc atacctgacc tccatagcag aaagtcaaaa gcctccgacc
     2581 ggaggctttt gacttgatcg gcacgtaaga ggttccaact ttcaccataa tgaaataaga
     2641 tcactaccgg gcgtattttt tgagttatcg agattttcag gagctaagga agctaaaatg
     2701 agccatattc aacgggaaac gtcttgcttg aagccgcgat taaattccaa catggatgct
     2761 gatttatatg ggtataaatg ggctcgcgat aatgtcgggc aatcaggtgc gacaatctat
     2821 cgattgtatg ggaagcccga tgcgccagag ttgtttctga aacatggcaa aggtagcgtt
     2881 gccaatgatg ttacagatga gatggtcagg ctaaactggc tgacggaatt tatgcctctt
     2941 ccgaccatca agcattttat ccgtactcct gatgatgcat ggttactcac cactgcgatc
     3001 ccagggaaaa cagcattcca ggtattagaa gaatatcctg attcaggtga aaatattgtt
     3061 gatgcgctgg cagtgttcct gcgccggttg cattcgattc ctgtttgtaa ttgtcctttt
     3121 aacggcgatc gcgtatttcg tctcgctcag gcgcaatcac gaatgaataa cggtttggtt
     3181 ggtgcgagtg attttgatga cgagcgtaat ggctggcctg ttgaacaagt ctggaaagaa
     3241 atgcataaac tcttgccatt ctcaccggat tcagtcgtca ctcatggtga tttctcactt
     3301 gataacctta tttttgacga ggggaaatta ataggttgta ttgatgttgg acgagtcgga
     3361 atcgcagacc gataccagga tcttgccatc ctatggaact gcctcggtga gttttctcct
     3421 tcattacaga aacggctttt tcaaaaatat ggtattgata atcctgatat gaataaattg
     3481 cagtttcact tgatgctcga tgagtttttc taatgaggac ctaaatgtaa tcacctggct
     3541 caccttcggg tgggcctttc tgcgttgctg gcgtttttcc ataggctccg cccccctgac
     3601 gagcatcaca aaaatcgatg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     3661 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     3721 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     3781 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     3841 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     3901 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     3961 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     4021 gtatttggta tctgcgctct gctgaagcca gttacctcgg aaaaagagtt ggtagctctt
     4081 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     4141 cgcgcagaaa aaaaggatct caagaagatc ctttgatttt ctaccgaaga aaggccca
//
