LOCUS       Exported                6325 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pWW3561, complete sequence.
ACCESSION   KY776519
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6325)
  AUTHORS   Whitaker WR, Shepherd ES, Sonnenburg JL.
  TITLE     Tunable expression tools enable single-cell strain distinction in 
            the gut microbiome
  JOURNAL   Cell (2017) In press
REFERENCE   2  (bases 1 to 6325)
  AUTHORS   Whitaker WR, Shepherd ES, Sonnenburg JL.
  TITLE     Direct Submission
  JOURNAL   Submitted (15-MAR-2017) Microbiology & Immunology, Stanford 
            University School of Medicine, 299 Campus Drive, Stanford, CA 
            94305-5124, USA
REFERENCE   3  (bases 1 to 6325)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6325
                     /organism="Cloning vector pWW3561"
                     /mol_type="other DNA"
                     /db_xref="taxon:1977345"
     rep_origin      25..798
     rep_origin      29..417
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     oriT            588..697
                     /note="incP origin of transfer"
     regulatory      841..933
                     /regulatory_class="promoter"
     terminator      1056..1127
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1143..1170
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_RNA        1288..1337
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     regulatory      1362..1384
                     /regulatory_class="ribosome_binding_site"
     CDS             1385..2200
                     /codon_start=1
                     /product="green fluorescent protein - 3xFLAG 6xHis tag"
                     /label=green fluorescent protein - 3xFLAG 6xHis tag
                     /protein_id="ARI71370.1"
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYKGSDYKDDDDKDYKDDDDKDYKDDDDKGHHHHHH"
     misc_feature    1385..2098
                     /label=GFP domain of fusion protein
                     /note="GFP domain of fusion protein"
     CDS             1385..2098
                     /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"
     misc_feature    2099..2197
                     /label=epitope tag - 3xFLAG 6xHis
                     /note="epitope tag - 3xFLAG 6xHis"
     CDS             2105..2176
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags"
                     /label=3xFLAG
                     /translation="DYKDDDDKDYKDDDDKDYKDDDDK"
     CDS             2180..2197
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      2220..2247
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     regulatory      2222..2251
                     /regulatory_class="terminator"
     CDS             complement(2479..3762)
                     /codon_start=1
                     /product="NBU integrase"
                     /label=NBU integrase
                     /protein_id="ARI71368.1"
                     /translation="MNIKRNIIFALESRKKNGVPIVENVPIRMRVIFASQRIEFTTGYR
                     IDVAKWDADKQRVKSGCTNKLKQSAAEINTDLLKYYAEIQNIFKEFEVQEVMPTTQQLK
                     EAFNMRMKDTSEEQPEEAPVSFWEVFDEFVKECGNQNNWTASTYEKFAAVRNHLKEFKE
                     DATFNYFNEFGLNEYVNFLRDTKDMRNSTIGKQMGFLKWFLRWSFKKGHHQNIAYDTFK
                     PKLKTTSKKVIFLTWDELNKLKDYQIPKDKQYLERVRDVFLFCCFTSLRYSDVRNLKRS
                     DVKSDHIEITTVKTADSLTIELNKYSKAILDKYKDIHFENYMALPVISNQKMNDYLKEL
                     GELAEINEPVRETYYKGNERIDEVTPKYALLSTHAGRRTFICNALALGIPAQVVMKWTG
                     HSDYKAMKPYIDIADDIKANAMNKFNQL"
     misc_feature    3922..3933
                     /label=integration site
                     /note="integration site"
     misc_feature    4201..4798
                     /label=split antibiotic resistance marker
                     /note="split antibiotic resistance marker"
     CDS             complement(4202..5062)
                     /codon_start=1
                     /product="AmpR"
                     /label=AmpR
                     /protein_id="ARI71369.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     misc_feature    4795..5062
                     /label=split antibiotic resistance marker
                     /note="split antibiotic resistance marker"
     CDS             5550..6284
                     /codon_start=1
                     /product="ErmG"
                     /label=ErmG
                     /protein_id="ARI71371.1"
                     /translation="MNKVNIKDSQNFITSKYHIEKIMNCISLDEKDNIFEIGAGKGHFT
                     AGLVKRCNFVTAIEIDSKLCEVTRNKLLNYPNYQIVNDDILKFTFPSHNPYKIFGSIPY
                     NISTNIIRKIVFESSATISYLIVEYGFAKMLLDTNRSLALLLMAEVDISILAKIPRYYF
                     HPKPKVDSTLIVLKRKPAKMAFKERKKYETFVMKWVNKEYEKLFTKNQFNKALKHARIY
                     DINNISFEQFVSLFNSYKIFNG"