LOCUS       sequence-251724-        4414 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes the cROSA26 homology landing pad with GFP dropout 
            destination vector as a type 0 part to be used in the MTK system.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4414)
  AUTHORS   Fonseca JP, Bonny AR, Kumar GR, Ng AH, Town J, Wu QC, Aslankoohi E, 
            Chen SY, Dods G, Harrigan P, Osimiri LC, Kistler AL, El-Samad H
  TITLE     A Toolkit for Rapid Modular Construction of Biological Circuits in 
            Mammalian Cells
  JOURNAL   ACS Synth Biol. 2019 Nov 15;8(11):2593-2606. doi: 
            10.1021/acssynbio.9b00322. Epub 2019 Nov 5.
  PUBMED    31686495
REFERENCE   2  (bases 1 to 4414)
  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: 
            "10.1021/acssynbio.9b00322"; journalName: "ACS Synth Biol"; date: 
            "2019-11-15- 15"; volume: "8"; issue: "11"; pages: "2593-2606"
COMMENT     Sequence Label: MTK0_028
FEATURES             Location/Qualifiers
     source          1..4414
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..34
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     RBS             341..352
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             359..1072
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      1084..1155
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1171..1198
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     protein_bind    1571..1604
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     CDS             complement(2251..3066)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(2251..3066)
                     /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="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     2974..2993
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3058..3077
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      3291..3879
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3780..3799
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(3995..4011)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3995..4011)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 gaagttccta ttctctagaa agtataggaa cttcctgatg agacgtggta gagccacaaa
       61 cagccggtac aagcaacgat ctccaggacc atctgaatca tgcgcggatg acacgaactc
      121 acgacggcga tcacagacat taacccacag tacagacact gcgacaacgt ggcaattcgt
      181 cgcaatacaa cggaaagtga aacgtgattt catgcgtcat tttgaacatt ttgtaaatct
      241 tatttaataa tgtgtgcggc aattcacatt taatttatga atgttttctt aacatcgcgg
      301 caactcaaga aacggcaggt tcggatctta gctactagag aaagaggaga aatactagat
      361 gcgtaaaggc gaagagctgt tcactggtgt cgtccctatt ctggtggaac tggatggtga
      421 tgtcaacggt cataagtttt ccgtgcgtgg cgagggtgaa ggtgacgcaa ctaatggtaa
      481 actgacgctg aagttcatct gtactactgg taaactgccg gttccttggc cgactctggt
      541 aacgacgctg acttatggtg ttcagtgctt tgctcgttat ccggaccata tgaagcagca
      601 tgacttcttc aagtccgcca tgccggaagg ctatgtgcag gaacgcacga tttcctttaa
      661 ggatgacggc acgtacaaaa cgcgtgcgga agtgaaattt gaaggcgata ccctggtaaa
      721 ccgcattgag ctgaaaggca ttgactttaa agaggacggc aatatcctgg gccataagct
      781 ggaatacaat tttaacagcc acaatgttta catcaccgcc gataaacaaa aaaatggcat
      841 taaagcgaat tttaaaattc gccacaacgt ggaggatggc agcgtgcagc tggctgatca
      901 ctaccagcaa aacactccaa tcggtgatgg tcctgttctg ctgccagaca atcactatct
      961 gagcacgcaa agcgttctgt ctaaagatcc gaacgagaaa cgcgatcata tggttctgct
     1021 ggagttcgta accgcagcgg gcatcacgca tggtatggat gaactgtaca aatgaccagg
     1081 catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat ctgttgtttg
     1141 tcggtgaacg ctctctacta gagtcacact ggctcacctt cgggtgggcc tttctgcgtt
     1201 tatagctgga aatctgctcg tcagtggtgc tcacactgac gaatcatgta cagatcatac
     1261 cgatgactgc ctggcgactc acaactaagc aagacagccg gaaccagcgc cggcgaacac
     1321 cactgcatat atggcatatc acaacagtcc acgtctcaag cagttacaga gatgttacga
     1381 accctgaaca ggttgactat taattgtgtc tgcttgatgt ggacaccagg tggcgctgga
     1441 catcagattt ggagaggcag ttgtctaggg aaccgggctc tgtgccagcg caggaggcag
     1501 gctggctctc ctattccagg gatgctcatc caggaaggaa aggttgcatg ctggacacac
     1561 taaccttaca gaagttccta ttctctagaa agtataggaa cttcgagtag atcgcggggg
     1621 agggagggat tcttctaggc ccagggcggt ccttaggaga caggaggcag ccgagaactc
     1681 ccataaaggt attgcggcac tcccctcccc ctaccgagaa gggtgcggcc ttctctccgc
     1741 ctcctccact gcagatccct caggattgca gctcgcgccg attttcagag atcacgcacc
     1801 tccctcccac aaaccagcag gaccgacccc cgctcctccc cccacctccg cgagtccgcg
     1861 agcggatcca gctcatctcg tccttcaggc ggcggacgcc cgggacggag ccgcggtcgc
     1921 cgccacccag aagcctcgac ccagcagtcc gcctccgcct ccgcctccgc ctctagtgcg
     1981 tgcgcagggg cgcggggcca gacactgcgg cgcggggcct cgggaaggac cgtaccaacg
     2041 accgcctccc tgcccctccg ccccctctgc tgacccgctc agtggtttcg ctcaattggg
     2101 acccgagcca atggcaaggg gccgttcccc ggcttcagca gccaatcagg tgccgaagcc
     2161 acacgggcgg cgggtaaccc tactccccgc gagggttttc ccagtcacga cccgagcggc
     2221 cgcgtgttac aaccaattaa ccaattctga ttagaaaaac tcatcgagca tcaaatgaaa
     2281 ctgcaattta ttcatatcag gattatcaat accatatttt tgaaaaagcc gtttctgtaa
     2341 tgaaggagaa aactcaccga ggcagttcca taggatggca agatcctggt atcggtctgc
     2401 gattccgact cgtccaacat caatacaacc tattaatttc ccctcgtcaa aaataaggtt
     2461 atcaagtgag aaatcaccat gagtgacgac tgaatccggt gagaatggca aaagcttatg
     2521 catttctttc cagacttgtt caacaggcca gccattacgc tcgtcatcaa aatcactcgc
     2581 atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg aggcgaaata cgcgatcgct
     2641 gttaaaagga caattacaaa caggaatcga atgcaaccgg cgcaggaaca ctgccagcgc
     2701 atcaacaata ttttcacctg aatcaggata ttcttctaat acctggaatg ctgttttccc
     2761 ggggatcgca gtggtgagta accatgcatc atcaggagta cggataaaat gcttgatggt
     2821 cggaagaggc ataaattccg tcagccagtt tagtctgacc atctcatctg taacatcatt
     2881 ggcaacgcta cctttgccat gtttcagaaa caactctggc gcatcgggct tcccatacaa
     2941 tcgatagatt gtcgcacctg attgcccgac attatcgcga gcccatttat acccatataa
     3001 atcagcatcc atgttggaat ttaatcgcgg cctggagcaa gacgtttccc gttgaatatg
     3061 gctcataaca ccccttgtat tactgtttat gtaagcagac agttttattg ttcatgatga
     3121 tatattttta tcttgtgcaa tgtaacatca gagattttga gacacaacgt ggctttgttg
     3181 aataaatcga acttttgctg agttgaagga tcagtcatga ccaaaatccc ttaacgtgag
     3241 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     3301 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     3361 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     3421 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     3481 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     3541 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     3601 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     3661 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     3721 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     3781 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     3841 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     3901 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     3961 gattctgtgg ataaccgtgc ggccgccaat catggtcata gctgtttcct gcaacggaaa
     4021 gggcctaaaa ttttaagatt agtatctaat ttaaaatgct acactttggt ggtgtaaaca
     4081 acgttctcaa acgacgtgag taaaaaactc agatgaccat gcccttaacc tccttttatc
     4141 cacaggaaaa gcactacaaa aacgatctta agtagtaata acacgcagcc ttgctgcata
     4201 aaatcagccc gagatgaatg cctggactcc catttccctc ctccacccct attataatta
     4261 ttttgtgaca caaactcaag cagggtgaag agagaaatct cacacctgct caatcccctg
     4321 caggacaacg cccagacacc aggtaaagcc tgggcacaga gagtcccgcc cacaatcgag
     4381 aaggagcaac tcgacatgga ggcgatgacg ccct
//
