LOCUS       sequence-213563-        3734 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  This plasmid is used for efficient cloning of DNA and 
            temperature-induced expression of protein in E. coli..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3734)
  AUTHORS   Gao H, Qi X, Hart DJ, Gao S, Wang H, Xu S, Zhang Y, Liu X, Liu Y, An
            Y
  TITLE     Three Novel Escherichia coli Vectors for Convenient and Efficient 
            Molecular Biological Manipulations.
  JOURNAL   J Agric Food Chem. 2018 Jun 20;66(24):6123-6131. doi: 
            10.1021/acs.jafc.8b01960. Epub 2018 Jun 11.
  PUBMED    29798665
REFERENCE   2  (bases 1 to 3734)
  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"; doi: 
            "10.1021/acs.jafc.8b01960"; journalName: "J Agric Food Chem"; date: 
            "2018-06-20- 20"; volume: "66"; issue: "24"; pages: "6123-6131"
COMMENT     Sequence Label: pANY3
FEATURES             Location/Qualifiers
     source          1..3734
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..18
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(66..84)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      80..127
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(195..200,201..217,218..223))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     CDS             complement(236..1051)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(236..1051)
                     /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
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     959..978
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     1043..1062
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      1173..1761
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1662..1681
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(1842..2555)
                     /codon_start=1
                     /gene="cIts"
                     /product="temperature-sensitive variant of the phage lambda
                     repressor"
                     /label=lambda repressor (ts)
                     /note="thermosensitivity is conferred by the A67T mutation"
                     /translation="MSTKKKPLTQEQLEDARRLKAIYEKKKNELGLSQESVADKMGMGQ
                     SGVGALFNGINALNAYNAALLTKILKVSVEEFSPSIAREIYEMYEAVSMQPSLRSEYEY
                     PVFSHVQAGMFSPKLRTFTKGDAERWVSTTKKASDSAFWLEVEGNSMTAPTGSKPSFPD
                     GMLILVDPEQAVEPGDFCIARLGGDEFTFKKLIRDSGQVFLQPLNPQYPMIPCNESCSV
                     VGKVIASQWPEETFG"
     RBS             2644..2652
                     /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)"
     protein_bind    2901..2915
                     /label=boxB
                     /bound_moiety="lambda-N peptide"
                     /note="lambda N peptide binding site consisting of an RNA 
                     hairpin within the N-utliization (nut) site of the lambda 
                     bacteriophage transcript (Tan and Frankel, 1995)"
     CDS             3354..3659
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     3538..3557
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
ORIGIN
        1 caccaccacc accaccacta aggctgctaa caaagcccga aaggaagctg agttggctgc
       61 tgccaccgct gagcaataac tagcataacc ccttggggcc tctaaacggg tcttgagggg
      121 ttttttgctg aaaggaggaa ctatatccgg atatccctga ctgcgttagc aatttaactg
      181 tgataaacta ccgcattaaa gcttatcgat gataagctgt caaacatgag aattcttaga
      241 aaaactcatc gagcatcaaa tgaaactgca atttattcat atcaggatta tcaataccat
      301 atttttgaaa aagccgtttc tgtaatgaag gagaaaactc accgaggcag ttccatagga
      361 tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc aacatcaata caacctatta
      421 atttcccctc gtcaaaaata aggttatcaa gtgagaaatc accatgagtg acgactgaat
      481 ccggtgagaa tggcaaaagt ttgtgcattt ctttccagac ttgttcaaca ggccagccat
      541 tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt attcattcgt gattgcgcct
      601 gagcgagacg aaatacgcga tcgctgttaa aaggacaatt acaaacagga atcgaatgca
      661 accggcgcag gaacactgcc agcgcatcaa caatattttc acctgaatca ggatattctt
      721 ctaatacctg gaatgctgtt ttcccgggga tcgcagtggt gagtaaccat gcgtcatcag
      781 gagtacggat aaaatgcttg atggtcggaa gaggcataaa ttccgtcagc cagtttagtc
      841 tgaccatctc atctgtaaca tcattggcaa cgctaccttt gccatgtttc agaaacaact
      901 ctggcgcatc gggcttccca tacaatcgat agattgtcgc acctgattgc ccgacattat
      961 cgcgagccca tttataccca tataaatcag catccatgtt ggaatttaat cgcggcctcg
     1021 agcaagacgt ttcccgttga atatggctca taacacccct tgtattactg tttatgtaag
     1081 cagacagttt tattgttcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     1141 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     1201 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     1261 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc
     1321 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     1381 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     1441 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     1501 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     1561 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     1621 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     1681 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     1741 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     1801 tgctggcctt tggtaccgcg ccgaccagaa caccttgccg atcagccaaa cgtctcttca
     1861 ggccactgac tagcgataac tttccccaca acggaacaac tctcattgca tgggatcatt
     1921 gggtactgtg ggtttagtgg ttgtaaaaac acctgaccgc tatccctgat cagtttcttg
     1981 aaggtaaact catcaccccc aagtctggct atgcagaaat cacctggctc aacagcctgc
     2041 tcagggtcaa cgagaattaa cattccgtca ggaaagcttg gcttggagcc tgttggtgcg
     2101 gtcatggaat taccttcaac ctcaagccag aatgcagaat cactggcttt tttggttgtg
     2161 cttacccatc tctccgcatc acctttggta aaggttctaa gcttaggtga gaacatccct
     2221 gcctgaacat gagaaaaaac agggtactca tactcacttc taagtgacgg ctgcatacta
     2281 accgcttcat acatctcgta gatttctctg gcgattgaag ggctaaattc ttcaacgcta
     2341 actttgagaa tttttgtaag caatgcggcg ttataagcat ttaatgcatt gatgccatta
     2401 aataaagcac caacgcctga ctgccccatc cccatcttgt ctgcgacaga ttcctgggat
     2461 aagccaagtt catttttctt tttttcataa attgctttaa ggcgacgtgc gtcctcaagc
     2521 tgctcttgtg ttaatggttt cttttttgtg ctcatacgtt aaatctatca ccgcaaggga
     2581 taaatatcta acaccgtgcg tgttgactat tttacctctg gcggtgataa tggttgcatg
     2641 tactaaggag gttgtatgga acaacgcata accctgaaag attatgcaat gcgctttggg
     2701 caaaccaaga cagctaaaga tctctcacct accaaacaat gcccccctgc aaaaaataaa
     2761 ttcatataaa aaacatacag ataaccatct gcggtgataa attatctctg gcggtgttga
     2821 cataaatacc actggcggtg atactgagca catcagcagg acgcactgac caccatgaag
     2881 gtgacgctct taaaaattaa gccctgaaga agggcagcat tcaaagcaga aggctttggg
     2941 gtgtgtgata cgaaacgaag cattggttaa aaattaagga ggaacatatg actagtaggc
     3001 ctgtcgaccc tgggtctgga gaccggctta ctaaaagcca gataacagta tgcgtatttg
     3061 cgcgctgatt tttgcggtat aagaatatat actgatatgt atacccgaag tatgtcaaaa
     3121 agaggtatgc tatgaagcag cgtattacag tgacagttga cagcgacagc tatcagttgc
     3181 tcaaggcata tatgatgtca atatctccgg tctggtaagc acaaccatgc agaatgaagc
     3241 ccgtcgtctg cgtgccgaac gctggaaagc ggaaaatcag gaagggatgg ctgaggtcgc
     3301 ccggtttatt gaaatgaacg gctcttttgc tgacgagaac aggggctggt gaaatgcagt
     3361 ttaaggttta cacctataaa agagagagcc gttatcgtct gtttgtggat gtacagagtg
     3421 atattattga cacgcccggg cgacggatgg tgatccccct ggccagtgca cgtctgctgt
     3481 cagataaagt ctcccgtgaa ctttacccgg tggtgcatat cggggatgaa agctggcgca
     3541 tgatgaccac cgatatggcc agtgtgccgg tctccgttat cggggaagaa gtggctgatc
     3601 tcagccaccg cgaaaatgac atcaaaaacg ccattaacct gatgttctgg ggaatataaa
     3661 tgtcaggctc ccttatacac agccagtcgg tctctcgtcg acctatggtc aggcctggat
     3721 ccgagctcct gcag
//
