LOCUS       Exported                8737 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Gateway recycling vector pRED56-MD8-Pnos-42 DNA, complete sequence.
ACCESSION   LC094300
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8737)
  AUTHORS   Saisho T, Shibahara K, Nakao A, Tanaka Y, Nishimura K, Kimura T, 
            Nakagawa T.
  TITLE     Improved Gateway Recycling Cloning System for Multigene Construction
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8737)
  AUTHORS   Saisho T, Shibahara K, Nakao A, Tanaka Y, Nishimura K, Kimura T, 
            Nakagawa T.
  TITLE     Direct Submission
  JOURNAL   Submitted (02-NOV-2015) Contact:Tsuyoshi Nakagawa Shimane 
            University, Interdisciplinary Center for Science Research; 1060 
            Nishikawatsu, Matsue, Shimane 690-8504, Japan
REFERENCE   3  (bases 1 to 8737)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     constructed using pDONR201.
FEATURES             Location/Qualifiers
     source          1..8737
                     /organism="Gateway recycling vector pRED56-MD8-Pnos-42"
                     /mol_type="other DNA"
                     /db_xref="taxon:1747861"
     misc_feature    complement(28..127)
                     /label=attL5
                     /note="attL5"
     protein_bind    29..124
                     /gene="mutant version of attL"
                     /label=attL5
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     misc_feature    674..1071
                     /label=MD8
                     /note="MD8"
     regulatory      1078..1380
                     /regulatory_class="promoter"
                     /note="NOS promoter"
     promoter        1160..1339
                     /label=NOS promoter
                     /note="nopaline synthase promoter"
     promoter        1192..1375
                     /function="NOS promoter"
                     /label=NOS promoter
                     /note="nopaline synthase promoter"
     misc_feature    1393..2109
                     /label=yellow fluorescent protein
                     /note="yellow fluorescent protein"
     CDS             1393..2109
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     misc_feature    complement(2119..2243)
                     /label=attR1
                     /note="attR1"
     protein_bind    2119..2243
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        2280..2310
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             2364..3023
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=cat
                     /protein_id="BAT33472.1"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     gene            2364..3023
                     /gene="cat"
                     /label=cat
     CDS             3365..3670
                     /codon_start=1
                     /gene="ccdB"
                     /label=ccdB
                     /protein_id="BAT33473.1"
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     gene            3365..3670
                     /gene="ccdB"
                     /label=ccdB
     misc_feature    3711..3835
                     /label=attR2
                     /note="attR2"
     protein_bind    complement(3711..3835)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     regulatory      3851..4115
                     /regulatory_class="terminator"
                     /note="NOS terminator"
     terminator      3861..4113
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     misc_feature    complement(4322..4446)
                     /label=attR4
                     /note="attR4"
     protein_bind    4322..4446
                     /gene="mutant version of attR"
                     /label=attR4
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        4471..4501
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     misc_feature    4773..4813
                     /label=rare cutter sites
                     /note="rare cutter sites"
     CDS             complement(4902..5762)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=bla
                     /note="ampicillin resistance"
                     /protein_id="BAT33474.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            complement(4902..5762)
                     /gene="bla"
                     /label=bla
     promoter        complement(5763..5867)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    6164..6288
                     /label=attR3
                     /note="attR3"
     protein_bind    complement(6164..6287)
                     /gene="mutant version of attR"
                     /label=attR3
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     misc_feature    6610..6709
                     /label=attL6
                     /note="attL6"
     protein_bind    complement(6613..6708)
                     /gene="mutant version of attL"
                     /label=attL6
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             6832..7641
                     /codon_start=1
                     /gene="kan"
                     /product="kanamycin resistance protein"
                     /label=kan
                     /protein_id="BAT33475.1"
                     /translation="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     gene            6832..7641
                     /gene="kan"
                     /label=kan
     rep_origin      7762..8401
                     /label=pUC origin
                     /note="pUC origin"
     rep_origin      7784..8372
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      8507..8534
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      8626..8712
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"