LOCUS       sequence-265179-        2985 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Low Phosphate inducible Nef.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2985)
  AUTHORS   Menacho-Melgar R, Ye Z, Moreb EA, Yang T, Efromson JP, Decker JS, 
            Wang R, Lynch MD
  TITLE     Scalable, two-stage, autoinduction of recombinant protein expression
            in E. coli utilizing phosphate depletion.
  JOURNAL   Biotechnol Bioeng. 2020 May 22. doi: 10.1002/bit.27440.
  PUBMED    32441815
REFERENCE   2  (bases 1 to 2985)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Biotechnol 
            Bioeng. 2020 May 22. doi: 10.1002/bit.27440."
COMMENT     Sequence Label: pHCKan-INSyibp-Nef
FEATURES             Location/Qualifiers
     source          1..2985
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      65..96
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     promoter        97..199
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             200..1015
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNGDRVFRLAQAQSRMNNGLVGA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(273..292)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      1039..1066
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     rep_origin      complement(1078..1665)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1158..1177)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(1687..1716)
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     terminator      1800..1871
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1887..1914
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     RBS             2176..2187
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             2825..2848
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             2849..2866
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
ORIGIN
        1 gacgaattct ctagatatcg ctcaatactg accatttaaa tcatacctga cctccatagc
       61 agaaagtcaa aagcctccga ccggaggctt ttgacttgat cggcacgtaa gaggttccaa
      121 ctttcaccat aatgaaataa gatcactacc gggcgtattt tttgagttat cgagattttc
      181 aggagctaag gaagctaaaa tgagccatat tcaacgggaa acgtcttgct cgaggccgcg
      241 attaaattcc aacatggatg ctgatttata tgggtataaa tgggctcgcg ataatgtcgg
      301 gcaatcaggt gcgacaatct atcgattgta tgggaagccc gatgcgccag agttgtttct
      361 gaaacatggc aaaggtagcg ttgccaatga tgttacagat gagatggtca ggctaaactg
      421 gctgacggaa tttatgcctc ttccgaccat caagcatttt atccgtactc ctgatgatgc
      481 atggttactc accactgcga tcccagggaa aacagcattc caggtattag aagaatatcc
      541 tgattcaggt gaaaatattg ttgatgcgct ggcagtgttc ctgcgccggt tgcattcgat
      601 tcctgtttgt aattgtcctt ttaacggcga tcgcgtattt cgtctcgctc aggcgcaatc
      661 acgaatgaat aacggtttgg ttggtgcgag tgattttgat gacgagcgta atggctggcc
      721 tgttgaacaa gtctggaaag aaatgcataa gcttttgcca ttctcaccgg attcagtcgt
      781 cactcatggt gatttctcac ttgataacct tatttttgac gaggggaaat taataggttg
      841 tattgatgtt ggacgagtcg gaatcgcaga ccgataccag gatcttgcca tcctatggaa
      901 ctgcctcggt gagttttctc cttcattaca gaaacggctt tttcaaaaat atggtattga
      961 taatcctgat atgaataaat tgcagtttca cttgatgctc gatgagtttt tctaatgagg
     1021 gcccaaatgt aatcacctgg ctcaccttcg ggtgggcctt tctgcgttgc tggcgttttt
     1081 ccataggctc cgcccccctg acgagcatca caaaaatcga tgctcaagtc agaggtggcg
     1141 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     1201 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     1261 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     1321 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     1381 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     1441 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     1501 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctc
     1561 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     1621 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatt
     1681 ttctaccgaa gaaaggccca cccgtgaagg tgagccagtg agttgattgc agtccagtta
     1741 cgctggagtc tgaggctcgt cctgaatgat atcaagcttg aattcgtttg cccaggcatc
     1801 aaataaaacg aaaggctcag tcgaaagact gggcctttcg ttttatctgt tgtttgtcgg
     1861 tgaacgctct ctactagagt cacactggct caccttcggg tgggcctttc tgcgtttata
     1921 cacagctaac accacgtcgt ccctatctgc tgccctaggt ctatgagtgg ttgctggata
     1981 acgtgcgtaa ttgtgctgat ctcttatata gctgctctca ttatctctct accctgaagt
     2041 gactctctca cctgtaaaaa taatatctca caggcttaat agtttcttaa tacaaagcct
     2101 gtaaaacgtc aggataactt ctatattcag ggagaccaca acggtttccc tctacaaata
     2161 attttgttta actttaaaga ggagaaatac tagaatgggt ggcaagtggt caaaacgtag
     2221 tatgggtgga tggtctgcta taagggaaag aatgagacga gctgagccac gagctgagcc
     2281 agcagcagat ggggtgggag cagtatctcg agacctggaa aaacatggag caatcacaag
     2341 tagcaataca gcagctacta atgctgattg tgcctggcta gaagcacaag aggaggaaga
     2401 ggtgggtttt ccagtcagac ctcaggtacc tttaagacca atgacttaca aggcagcttt
     2461 agatattagc cactttttaa aagaaaaggg gggactggaa gggctaattt ggtcccaaag
     2521 aagacaagag atccttgatc tgtggatcta ccacacacaa ggctacttcc ctgattggca
     2581 gaattacaca ccagggccag ggatcagata tccactgacc tttggatggt gcttcaagct
     2641 agtaccagtt gagccagaga aggtagaaga ggccaatgaa ggagagaaca acagcttgtt
     2701 acaccctatg agcctgcatg ggatggagga cgcggagaaa gaagtgttag tgtggaggtt
     2761 tgacagcaaa ctagcatttc atcacatggc ccgagagctg catccggagt actacaaaga
     2821 ctgcctcgaa gttctgttcc aggggcccca tcaccatcat caccactaat gaccggctta
     2881 tcggtcagtt tcacctgatt tacgtaaaaa cccgcttcgg cgggtttttg cttttggagg
     2941 ggcagaaaga tgaatgactg tccacgacgc tatacccaaa agaaa
//
