LOCUS       sequence-178163-        4008 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  MultiSite Gateway entry clones, second fragment (attL5, attL2) Human
            SMAD3 CDS.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4008)
  AUTHORS   Jackel C, Nogueira MS, Ehni N, Kraus C, Ranke J, Dohmann M, Noessner
            E, Nelson PJ
  TITLE     A vector platform for the rapid and efficient engineering of stable 
            complex transgenes.
  JOURNAL   Sci Rep. 2016 Oct 3;6:34365. doi: 10.1038/srep34365.
  PUBMED    27694838
REFERENCE   2  (bases 1 to 4008)
  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: "Sci Rep."; 
            date: "2016-10-3"; volume: "6:34365. doi"; pages: " 
            10.1038/srep34365"
COMMENT     Sequence Label: pENTR221-SMAD3
FEATURES             Location/Qualifiers
     source          1..4008
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     92..109
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(268..295)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(387..473)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     522..544
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     536..553
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     537..553
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    570..665
                     /gene="mutant version of attL"
                     /label=attL5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             708..731
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     protein_bind    complement(2120..2219)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(2249..2268)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2250..2268)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2273..2289)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2273..2289)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2391..2410)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2402..3211
                     /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="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2469..2488)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      3358..3946
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3847..3866
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
       61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
      121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
      181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc
      241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
      301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
      361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
      421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg
      481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa
      541 aacgacggcc agtcttaagc tcgggcccca aataatgatt ttattttgac tgatagtgac
      601 ctgttcgttg caacaaattg atgagcaatg cttttttata atgccaactt tgtatacaaa
      661 agttgtacag agctcgttta gtgaaccgtc agaattgatc taccatggac tacaaagacg
      721 atgacgacaa gcttgcggcc gcgaattcca tgtcgtccat cttgccattc actcccccga
      781 tcgtgaagcg cctgctgggc tggaagaagg gcgagcagaa cgggcaggag gagaaatggt
      841 gcgagaaggc ggtcaagagc ctggtcaaga aactcaagaa gacggggcag ctggacgagc
      901 tggagaaggc catcaccacg cagaacgtca acaccaagtg catcaccatc cccaggtccc
      961 tggatggccg gttgcaggtg tcccatcgga aggggctccc tcatgtcatc tactgccgcc
     1021 tgtggcgatg gccagacctg cacagccacc acgagctacg ggccatggag ctgtgtgagt
     1081 tcgccttcaa tatgaagaag gacgaggtct gcgtgaatcc ctaccactac cagagagtag
     1141 agacaccagt tctacctcct gtgttggtgc cacgccacac agagatcccg gccgagttcc
     1201 ccccactgga cgactacagc cattccatcc ccgaaaacac taacttcccc gcaggcatcg
     1261 agccccagag caatattcca gagaccccac cccctggcta cctgagtgaa gatggagaaa
     1321 ccagtgacca ccagatgaac cacagcatgg acgcaggttc tccaaaccta tccccgaatc
     1381 cgatgtcccc agcacataat aacttggacc tgcagccagt tacctactgc gagccggcct
     1441 tctggtgctc catctcctac tacgagctga accagcgcgt cggggagaca ttccacgcct
     1501 cgcagccatc catgactgtg gatggcttca ccgacccctc caattcggag cgcttctgcc
     1561 tagggctgct ctccaatgtc aacaggaatg cagcagtgga gctgacacgg agacacatcg
     1621 gaagaggcgt gcggctctac tacatcggag gggaggtctt cgcagagtgc ctcagtgaca
     1681 gcgctatttt tgtccagtct cccaactgta accagcgcta tggctggcac ccggccaccg
     1741 tctgcaagat cccaccagga tgcaacctga agatcttcaa caaccaggag ttcgctgccc
     1801 tcctggccca gtcggtcaac cagggctttg aggctgtcta ccagttgacc cgaatgtgca
     1861 ccatccgcat gagcttcgtc aaaggctggg gagcggagta caggagacag actgtgacca
     1921 gtaccccctg ctggattgag ctgcacctga atgggccttt gcagtggctt gacaaggtcc
     1981 tcacccagat gggctcccca agcatccgct gttccagtgt gtcttagaga gacatcaagt
     2041 gctctagagg atcccgggtg gcatccctgt gacccctccc cagtgcctct cctggccctg
     2101 gaagttgcca ctccagtgaa cccagctttc ttgtacaaag ttggcattat aagaaagcat
     2161 tgcttatcaa tttgttgcaa cgaacaggtc actatcagtc aaaataaaat cattatttgc
     2221 catccagctg cagggcggcc gcgatatccc ctatagtgag tcgtattaca tggtcatagc
     2281 tgtttcctgg cagctctggc ccgtgtctca aaatctctga tgttacattg cacaagataa
     2341 aaatatatca tcatgaacaa taaaactgtc tgcttacata aacagtaata caaggggtgt
     2401 tatgagccat attcaacggg aaacgtcgag gccgcgatta aattccaaca tggatgctga
     2461 tttatatggg tataaatggg ctcgcgataa tgtcgggcaa tcaggtgcga caatctatcg
     2521 cttgtatggg aagcccgatg cgccagagtt gtttctgaaa catggcaaag gtagcgttgc
     2581 caatgatgtt acagatgaga tggtcagact aaactggctg acggaattta tgcctcttcc
     2641 gaccatcaag cattttatcc gtactcctga tgatgcatgg ttactcacca ctgcgatccc
     2701 cggaaaaaca gcattccagg tattagaaga atatcctgat tcaggtgaaa atattgttga
     2761 tgcgctggca gtgttcctgc gccggttgca ttcgattcct gtttgtaatt gtccttttaa
     2821 cagcgatcgc gtatttcgtc tcgctcaggc gcaatcacga atgaataacg gtttggttga
     2881 tgcgagtgat tttgatgacg agcgtaatgg ctggcctgtt gaacaagtct ggaaagaaat
     2941 gcataaactt ttgccattct caccggattc agtcgtcact catggtgatt tctcacttga
     3001 taaccttatt tttgacgagg ggaaattaat aggttgtatt gatgttggac gagtcggaat
     3061 cgcagaccga taccaggatc ttgccatcct atggaactgc ctcggtgagt tttctccttc
     3121 attacagaaa cggctttttc aaaaatatgg tattgataat cctgatatga ataaattgca
     3181 gtttcatttg atgctcgatg agtttttcta atcagaattg gttaattggt tgtaacactg
     3241 gcagagcatt acgctgactt gacgggacgg cgcaagctca tgaccaaaat cccttaacgt
     3301 gagttacgcg tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     3361 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     3421 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     3481 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     3541 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     3601 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     3661 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     3721 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     3781 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     3841 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     3901 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     3961 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgtt
//
