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pToxin-mfLON-ecoRI vector (V002566#)

Basic Vector Information

Basic Vector Information
Vector Name pToxin-mfLON-ecoRI Length 5729 bp
Type Cloning vector Source Chan CT, Lee JW, Cameron DE, Bashor CJ, Collins JJ.

pToxin-mfLON-ecoRI vector Vector Map

pLscrO-2 RBS mfLON protease T0 pLscrO-2 RBS type-2 restriction enzyme EcoRI T1 p15A lambda t0 terminator beta-lactmase AmpR promoter

Plasmid Resuspension protocol:

1. Centrifuge at 5,000×g for 5 min.

2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.

3. Close the tube and incubate for 10 minutes at room temperature.

4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.

5.Store the plasmid at -20 ℃.

pToxin-mfLON-ecoRI vector Sequence

Copy Sequence

Download GeneBank File(.gb)

LOCUS       Exported                5729 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pToxin-mfLON-ecoRI, complete sequence.
ACCESSION   KT895277
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5729)
  AUTHORS   Chan CT, Lee JW, Cameron DE, Bashor CJ, Collins JJ.
  TITLE     'Deadman' and 'Passcode' microbial kill switches for bacterial 
            containment
  JOURNAL   Nat. Chem. Biol. 12 (2), 82-86 (2016)
  PUBMED    26641934
REFERENCE   2  (bases 1 to 5729)
  AUTHORS   Chan CTY., Lee JW, Cameron DE, Bashor CJ, Collins JJ.
  TITLE     Direct Submission
  JOURNAL   Submitted (12-OCT-2015) Department of Biological Engineering, 
            Massachusetts Institute of Technology, 77 Massachusetts Ave., 
            E25-302b, Cambridge, MA 02139, USA
REFERENCE   3  (bases 1 to 5729)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..5729
                     /organism="Cloning vector pToxin-mfLON-ecoRI"
                     /mol_type="other DNA"
                     /db_xref="taxon:1758747"
     regulatory      7..54
                     /regulatory_class="promoter"
                     /note="pLscrO-2"
     regulatory      81..101
                     /regulatory_class="ribosome_binding_site"
                     /note="RBS"
     CDS             103..2547
                     /codon_start=1
                     /product="mfLON protease"
                     /label=mfLON protease
                     /protein_id="ALP32113.1"
                     /translation="MSKKIKLPIFQIRGSFIVPGIKENLEVGRKNTLASVNYAIKNSNN
                     QMIAIPQIDASVEKPEFSDLHEFGILIDFEVIKEWKDNSLTISTNPIQRCKVISFFENE
                     DQVPYAEVELIESINDFSDEELKELIEKISDAIKTKASLVTKQIKQLISGESDDLSLAF
                     DSIMFKLAPSKILTNPEYITSPSLKTRWSIIEKIIFAEDGIITRNAESIDAARQKNEIE
                     QELNHKLKEKMDKQQKEYYLREKMRIIKDELEDEDDSDDSSLEKYKERLAKEPFPEEVK
                     RKIMASIKRVEALQSGTPEWNTEKNYIDWMMSIPWWEETEDLTDLKYAKKILDKHHYGM
                     KKVKERIIEYLAVKTKTKSLKAPIITLVGPPGVGKTSLAKSIAEAVGKNFVKVSLGGVK
                     DESEIRGHRKTYVGSMPGRIIQTMKRAKVKNPLFLLDEIDKMASDHRGDPASAMLEVLD
                     PEQNKEFSDHYIEEPYDLSQVMFIATANYPEDIPEALYDRMEIINLSSYTEIEKVKIAQ
                     DYLVPKAIEQHELTSEEISFTEGAINEIIKYYTREAGVRQLERHINSIIRKYIVKNLNG
                     EMDKIVIDEKQVNDLLGKRIFDHTEKQEESQIGVVTGLAYTQFGGDILPIEVSLYPGKG
                     NLILTGKLGEVMKESATIALTYVKSNFEKFGVDKKVFEENDIHVHVPEGAVPKDGPSAG
                     ITITTALISALSDKPVSKEIGMTGEITLRGNVLPIGGLREKSISASRSGLKTIIIPKKN
                     ERDLDEIPDEVKAKLKIIPAEKYEEVFAIVFKTKAANKNEENTNEVPTFMLNAGQANRR
                     RV"
     regulatory      2561..2655
                     /regulatory_class="terminator"
                     /note="T0"
     terminator      2561..2655
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     regulatory      2662..2709
                     /regulatory_class="promoter"
                     /note="pLscrO-2"
     regulatory      2742..2761
                     /regulatory_class="ribosome_binding_site"
                     /note="RBS"
     CDS             2768..3601
                     /codon_start=1
                     /product="type-2 restriction enzyme EcoRI"
                     /label=type-2 restriction enzyme EcoRI
                     /protein_id="ALP32114.1"
                     /translation="MSNKKQSNRLTEQHKLSQGVIGIFGDYAKAHDLAVGEVSKLVKKA
                     LSNEYPQLSFRYRDSIKKTEINEALKKIDPDLGGTLFVSNSSIKPDGGIVEVKDDYGEW
                     RVVLVAEAKHQGKDIINIRNGLLVGKRGDQDLMAAGNAIERSHKNISEIANFMLSESHF
                     PYVLFLEGSNFLTENISITRPDGRVVNLEYNSGILNRLDRLTAANYGMPINSNLCINKF
                     VNHKDKSIMLQAASIYTQGDGREWDSKIMFEIMFDISTTSLRVLGRDLFEQLTSK"
     regulatory      3621..3680
                     /regulatory_class="terminator"
                     /note="T1"
     terminator      3626..3712
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     rep_origin      3732..4528
                     /label=p15A
                     /note="p15A"
     rep_origin      3833..4378
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin."
     terminator      4540..4634
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(4664..5524)
                     /codon_start=1
                     /product="beta-lactmase"
                     /label=beta-lactmase
                     /protein_id="ALP32115.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(5525..5629)
                     /gene="bla"
                     /label=AmpR promoter

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