LOCUS       Exported                6218 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Expresses Br512 polymerase in E.coli.
ACCESSION   .
VERSION     .
KEYWORDS    pKAR2-Br512
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6218)
  AUTHORS   Maranhao A,Bhadra S, Paik I, Walker D, Ellington AD
  TITLE     An improved and readily available version of Bst DNA Polymerase for 
            LAMP, and applications to COVID-19 diagnostics
  JOURNAL   medRxiv
REFERENCE   2  (bases 1 to 6218)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "medRxiv"
FEATURES             Location/Qualifiers
     source          1..6218
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     terminator      16..62
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(359..985)
                     /codon_start=1
                     /gene="tetR"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     890..909
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     CDS             complement(1014..1874)
                     /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"
     primer_bind     1637..1656
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1875..1946)
                     /gene="bla"
                     /label=AmpR promoter
     terminator      2023..2109
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     complement(2253..2270)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2424..3012)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2504..2523)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3822..3841
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     3906..3925
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     4069..4087
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     protein_bind    4108..4126
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
     primer_bind     4127..4146
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4127..4145
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    4147..4165
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     misc_RNA        4185..4234
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     RBS             4268..4276
                     /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)"