LOCUS       Exported                6491 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pAP322, complete sequence.
ACCESSION   MF498902
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6491)
  AUTHORS   Park AS, Rubin EJ, Minch KJ, Sherman DR.
  TITLE     Clp-mediated Regulation of WhiB1 is Required for Cell Division in 
            Mycobacteria
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6491)
  AUTHORS   Park AS, Rubin EJ, Minch KJ, Sherman DR.
  TITLE     Direct Submission
  JOURNAL   Submitted (18-JUL-2017) Department of Immunology and Infectious 
            Diseases, Harvard School of Public Health, 665 Huntington Ave, 
            Boston, MA 02115, USA
REFERENCE   3  (bases 1 to 6491)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6491
                     /organism="Cloning vector pAP322"
                     /mol_type="other DNA"
                     /db_xref="taxon:2025120"
     regulatory      7..38
                     /regulatory_class="terminator"
     CDS             complement(70..693)
                     /codon_start=1
                     /product="TetR"
                     /label=TetR
                     /note="Tet Repressor class B"
                     /protein_id="ASU06031.1"
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     regulatory      complement(720..1036)
                     /regulatory_class="promoter"
     regulatory      complement(1055..1082)
                     /regulatory_class="terminator"
     terminator      1055..1082
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     rep_origin      1249..1868
     rep_origin      complement(1264..1852)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(2184..2999)
                     /codon_start=1
                     /product="aph"
                     /label=aph
                     /note="kanamycin resistance gene"
                     /protein_id="ASU06032.1"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPHAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHNLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(3280..4278)
                     /codon_start=1
                     /product="L5 integrase"
                     /label=L5 integrase
                     /protein_id="ASU06033.1"
                     /translation="MDAEAWLAGEKRLIEMETWTPPQDRAKKAAASAITLEEYTRKWLV
                     ERDLADGTRDLYSGHAERRIYPVLGEVAVTEMTPALVRAWWAGMGRKHPTARRHAYNVL
                     RAVMNTAVEDKLIAENPCRIEQKAADERDVEALTPEELDIVAAEIFEHYRIAAYILAWT
                     SLRFGELIELRRKDIVDDGMTMKLRVRRGASRVGNKIVVGNAKTVRSKRPVTVPPHVAE
                     MIRAHMKDRTKMNKGPEAFLVTTTQGNRLSKSAFTKSLKRGYAKIGRPELRIHDLRAVG
                     ATFAAQAGATTKELMARLGHTTPRMAMKYQMASEARDEAIAEAMSKLAKTS"
     terminator      4942..4988
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     regulatory      complement(4942..4985)
                     /regulatory_class="terminator"
     terminator      5034..5061
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     regulatory      5086..5391
                     /regulatory_class="promoter"
     protein_bind    5261..5278
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     regulatory      5407..5412
                     /regulatory_class="ribosome_binding_site"
     CDS             5420..6190
                     /codon_start=1
                     /product="his-eGFP"
                     /label=his-eGFP
                     /note="N-terminally his-tagged eGFP"
                     /protein_id="ASU06034.1"
                     /translation="MASKGEHHHHHHEFGTVSKGEELFTGVVPILVELDGDVNGHKFSV
                     SGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAM
                     PEGYVQERTIFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNS
                     HNVYIMADKQKNGIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSA
                     LSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGS"
     CDS             5438..5455
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             6191..6442
                     /codon_start=1
                     /product="WhiB1"
                     /label=WhiB1
                     /note="WhiB1 from Mycobacterium smegmatis str. MC2 155"
                     /protein_id="ASU06035.1"
                     /translation="DWRHKAVCRDEDPELFFPVGNSGPALAQIADAKLVCNRCPVTTEC
                     LSWALESGQDAGVWGGMSEDERRALKRRNARTKARTGV"