LOCUS       Exported               10199 bp ds-DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector pECL275, complete sequence.
ACCESSION   KM521209
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10199)
  AUTHORS   Cameron DE, Collins JJ.
  TITLE     Tunable protein degradation in bacteria
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 10199)
  AUTHORS   Cameron DE, Collins JJ.
  TITLE     Direct Submission
  JOURNAL   Submitted (12-SEP-2014) Biomedical Engineering, Boston University, 
            44 Cummington St., Boston, MA 02215, USA
REFERENCE   3  (bases 1 to 10199)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..10199
                     /organism="Cloning vector pECL275"
                     /mol_type="other DNA"
                     /db_xref="taxon:1565073"
     misc_feature    7..425
                     /note="lacZ; 5 region"
     primer_bind     complement(24..40)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      446..490
                     /regulatory_class="terminator"
                     /note="T3 terminator"
     terminator      455..482
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     regulatory      604..677
                     /regulatory_class="promoter"
                     /note="PLtetO promoter"
     promoter        604..677
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    604..622
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    629..647
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             688..696
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3 end of the 16S rRNA (Chen et al., 1994)"
     CDS             700..3063
                     /codon_start=1
                     /product="mf-lon"
                     /label=mf-lon
                     /protein_id="AIU94520.1"
                     /translation="MSKKIKLPIFQIRGSFIVPGIKENLEVGRKNTLASVNYAIKNSNN
                     QMIAIPQIDASVEKPEFSDLHEFGILIDFEVIKEWKDNSLTISTNPIQRCKVISFFENE
                     DQVPYAEVELIESINDFSDEELKELIEKISDAIKTKASLVTKQIKQLISGESDDLSLAF
                     DSIMFKLAPSKILTNPEYITSPSLKTRWSIIEKIIFAEDGIITRNAESIDAARQKNEIE
                     QELNHKLKEKMDKQQKEYYLREKMRIIKDELEDEDDSDDSSLEKYKERLAKEPFPEEVK
                     RKIMASIKRVEALQSGTPEWNTEKNYIDWMMSIPWWEETEDLTDLKYAKKILDKHHYGM
                     KKVKERIIEYLAVKTKTKSLKAPIITLVGPPGVGKTSLAKSIAEAVGKNFVKVSLGGVK
                     DESEIRGHRKTYVGSMPGRIIQTMKRAKVKNPLFLLDEIDKMASDHRGDPASAMLEVLD
                     PEQNKEFSDHYIEEPYDLSQVMFIATANYPEDIPEALYDRMEIINLSSYTEIEKVKIAQ
                     DYLVPKAIEQHELTSEEISFTEGAINEIIKYYTREAGVRQLERHINSIIRKYIVKNLNG
                     EMDKIVIDEKQVNDLLGKRIFDHTEKQEESQIGVVTGLAYTQFGGDILPIEVSLYPGKG
                     NLILTGKLGEVMKESATIALTYVKSNFEKFGVDKKVFEENDIHVHVPEGAVPKDGPSAG
                     ITITTALISALSDKPVSKEIGMTGEITLRGNVLPIGGLREKSISASRSGLKTIIIPKKN
                     ERDLDEIPDEVKAKLKIIPAEKYEEVFAIVFKTK"
     terminator      3106..3192
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     regulatory      3109..3153
                     /regulatory_class="terminator"
                     /note="T1 terminator"
     misc_feature    3208..3621
                     /note="lacZ; 3 region"
     rep_origin      complement(3919..4313)
                     /direction=LEFT
                     /label=R6K origin
                     /note="R6K origin"
     rep_origin      3922..4310
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     promoter        4420..4524
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4525..5385
                     /codon_start=1
                     /product="bla"
                     /label=bla
                     /note="ampicillin resistance"
                     /protein_id="AIU94521.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     CDS             complement(6952..8373)
                     /codon_start=1
                     /product="sacB"
                     /label=sacB
                     /protein_id="AIU94522.1"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     promoter        complement(8374..8819)
                     /gene="sacR"
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     misc_feature    9851..10094
                     /note="oriT; RP4"
     oriT            9986..10095
                     /note="incP origin of transfer"