LOCUS       Exported               10198 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pRR22, complete sequence.
ACCESSION   JN201958
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10198)
  AUTHORS   Ramsden R, Arms L, Davis TN, Muller EG.
  TITLE     An intein with genetically selectable markers provides a new 
            approach to internally label proteins with GFP
  JOURNAL   BMC Biotechnol. 11 (1), 71 (2011)
  PUBMED    21708017
REFERENCE   2  (bases 1 to 10198)
  AUTHORS   Ramsden R, Arms L, Davis TN, Muller EGD.
  TITLE     Direct Submission
  JOURNAL   Submitted (30-JUN-2011) Biochemistry, University of Washington, 1705
            NE Pacific St, HSB J461, Seattle, WA 98195, USA
REFERENCE   3  (bases 1 to 10198)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..10198
                     /organism="Cloning vector pRR22"
                     /mol_type="other DNA"
                     /db_xref="taxon:1055401"
     promoter        14..678
                     /gene="GAL1 and GAL10 genes of S. cerevisiae"
                     /label=GAL1,10 promoter
                     /note="divergent inducible promoter, regulated by Gal4"
     regulatory      65..679
                     /regulatory_class="promoter"
                     /note="GAL10-GAL1"
     protein_bind    230..347
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     CDS             688..2901
                     /codon_start=1
                     /product="NLS-GFP-(Pch PRP8 intein with G418r)-FLAG"
                     /label=NLS-GFP-
                     /protein_id="AEK22021.1"
                     /translation="MVPKKKRKVLIDPKKKRKVDPKKKRKVLIKGSKGEELFTGVVPIL
                     VELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTFTYGVQCFSRY
                     PDHMKRHDFFKSAMPEGYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKED
                     GNILGHKLEYNYNSHNVYIMADKQKNGIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGP
                     VLLPDNHYLSTQSALSKDPNEKRDHMVLLEFVTAAGITHGMDELYKGAGAGAGAGAGFW
                     EKACLAKGTRLLRCDGTEINVEDVREGDLLLGPDGEPRRAFNIVNGIDRLYRIKIGGEK
                     EDLVVTPNHILVLYREDGSKNVEKQTVEITAAEFAALSTEERSLYSAFTSPRAEKGAAG
                     AGAGAASGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKPDAPELFLK
                     HGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTAFQVLEEYP
                     DSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASDFDDERNGW
                     PVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIADRYQDLAIL
                     WNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFFGTAGAGAGASAQTHSFKIEQ
                     VSLESEKTEWAGFRVDKDQLYLRHDYLVLHNSGFEDYKDDDDK"
     CDS             694..714
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             724..744
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             748..768
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             781..1491
                     /codon_start=1
                     /product="Aequoria victoria green fluorescent protein"
                     /label=GFP
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTFTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     misc_feature    1540..2862
                     /label=Pch PRP8 intein with G418r
                     /note="Pch PRP8 intein with G418r"
     CDS             1906..2709
                     /codon_start=1
                     /gene="aph(3)-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="GKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKPD
                     APELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTAF
                     QVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASDF
                     DDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIADR
                     YQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             2875..2898
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        3693..3908
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             3909..4712
                     /codon_start=1
                     /product="ScURA3"
                     /label=ScURA3
                     /protein_id="AEK22022.1"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     misc_feature    6057..7906
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN4 centromere fused to the 
                     autonomously replicating sequence ARS1/ARS416"
     rep_origin      7453..7906
                     /label=ARS1
                     /note="S. cerevisiae autonomously replicating sequence 
                     ARS1"
     misc_feature    8048..8188
                     /label=bom
                     /note="basis of mobility region from pBR322"
     rep_origin      complement(8374..8962)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(9133..9993)
                     /codon_start=1
                     /product="ampicillin"
                     /label=ampicillin
                     /protein_id="AEK22023.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(9994..10098)
                     /gene="bla"
                     /label=AmpR promoter