LOCUS       Exported               11032 bp ds-DNA     circular SYN 17-DEC-2018
DEFINITION  S. pombe expression vector pDUAL-GFH31c DNA, complete sequence.
ACCESSION   AB364148
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11032)
  AUTHORS   Matsuyama A, Shirai A, Yashiroda Y, Kamata A, Horinouchi S, Yoshida 
            M.
  TITLE     pDUAL, a multipurpose, multicopy vector capable of chromosomal 
            integration in fission yeast
  JOURNAL   Yeast 21 (15), 1289-1305 (2004)
  PUBMED    15546162
REFERENCE   2  (bases 1 to 11032)
  AUTHORS   Matsuyama A, Shirai A, Yoshida M.
  TITLE     A series of promoters for constitutive expression of heterologous 
            genes in fission yeast
  JOURNAL   Yeast 25 (5), 371-376 (2008)
  PUBMED    18437702
REFERENCE   3  (bases 1 to 11032)
  AUTHORS   Matsuyama A, Shirai A, Yoshida M.
  TITLE     Direct Submission
  JOURNAL   Submitted (15-OCT-2007) Contact:Akihisa Matsuyama RIKEN, Chemical 
            Genetics Laboratory; Hirosawa 2-1, Wako, Saitama 351-0198, Japan URL
            :http://cgl.riken.go.jp/
REFERENCE   4  (bases 1 to 11032)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     Fission yeast expression vector.
FEATURES             Location/Qualifiers
     source          1..11032
                     /organism="Schizosaccharomyces pombe expression vector
                     pDUAL-GFH31c"
                     /mol_type="other DNA"
                     /db_xref="taxon:477426"
     regulatory      1..237
                     /regulatory_class="terminator"
                     /gene="ADH1"
     misc_feature    1..237
                     /gene="ADH1"
                     /label=derived from Saccharomyces cerevisiae
                     /note="derived from Saccharomyces cerevisiae"
     gene            1..237
                     /gene="ADH1"
                     /label=ADH1
     terminator      21..208
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     misc_feature    244..4768
                     /label=derived from Schizosaccharomyces pombe
                     /note="derived from Schizosaccharomyces pombe"
     gene            join(244..1470,4415..4768)
                     /pseudo
                     /gene="leu1"
                     /label=leu1
     regulatory      244..954
                     /regulatory_class="promoter"
                     /pseudo
                     /gene="leu1"
     CDS             join(955..1470,4415..4768)
                     /pseudo
                     /gene="leu1"
                     /label=leu1
     regulatory      complement(1483..1871)
                     /regulatory_class="terminator"
                     /gene="ura4"
     gene            complement(1483..1871)
                     /gene="ura4"
                     /label=ura4
     CDS             complement(1872..2666)
                     /codon_start=1
                     /gene="ura4"
                     /product="orotidine 5-phosphate decarboxylase"
                     /label=ura4
                     /note="internal EcoRV site is eliminated."
                     /protein_id="BAF90981.1"
                     /translation="MDARVFQSYSARAEGMKNPIAKELLALMEEKQSNLSVAVDLTKKS
                     EILELVDKIGPYVCVIKTHIDVVEDFDQDMVEKLVALGKKHRFLIFEDRKFADIGNTVK
                     LQYASGVYKIASWAHITNCHTVPGEGIIQGLKEVGLPLGRGLLLLAEMSSKGSLATGSY
                     TEKTLEWFEKHTDFCFGFIAGRRFPNLQSDYITMSPGIGLDVKGDGLGQQYRTPEEVIV
                     NCGSDIIIVGRGVYGAGRNPVVEAKRYREAGWKAYQQRLSQH"
     gene            complement(1872..2666)
                     /gene="ura4"
                     /label=ura4
     regulatory      complement(2667..3190)
                     /regulatory_class="promoter"
                     /gene="ura4"
     gene            complement(2667..3190)
                     /gene="ura4"
                     /label=ura4
     rep_origin      3208..4400
                     /standard_name="ars1"
                     /label=ars1
     rep_origin      3275..4058
                     /label=ars1
                     /note="Schizosaccharomyces pombe autonomously replicating 
                     sequence ars1"
     misc_feature    4769..7897
                     /label=derived from Cloning vector pUC119
                     /note="derived from Cloning vector pUC119"
     primer_bind     complement(4781..4797)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      5010..5465
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        5747..5851
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5852..6712
                     /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"
     rep_origin      6883..7471
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    7759..7780
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        7795..7825
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7833..7849
                     /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)."
     primer_bind     7857..7873
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      7898..8502
                     /regulatory_class="promoter"
                     /gene="cam1"
     misc_feature    7898..8502
                     /gene="cam1"
                     /label=derived from Schizosaccharomyces pombe
                     /note="derived from Schizosaccharomyces pombe"
     gene            7898..8502
                     /gene="cam1"
                     /label=cam1
     misc_feature    8531..10243
                     /standard_name="ccdB cassette reading frame B"
                     /label=ccdB cassette reading frame B
     protein_bind    8535..8659
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        8684..8714
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             8768..9427
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     CDS             9769..10074
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     protein_bind    complement(10115..10239)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             10247..10960
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=green fluorescent protein
                     /note="Ser65 is substituted by Cys."
                     /protein_id="BAF90982.1"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFCYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             10964..11011
                     /codon_start=1
                     /product="FLAG epitope and hexahistidine"
                     /label=FLAG epitope and hexahistidine
                     /protein_id="BAF90983.1"
                     /translation="MDYKDDDDKHHHHHH"
     CDS             10967..10990
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             10991..11008
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"