LOCUS       pKF_18k-2.        2204 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Bacterial cloning and site-directed mutagenesis vector with a 
            kanamycin resistance gene containing amber stop codons. The MCS is 
            reversed in pKF 19k-2.
ACCESSION   .
VERSION     .
KEYWORDS    pKF 18k-2
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 2204)
  AUTHORS   Hashimoto-Gotoh T, Yasojima K, Tsujimura A.
  TITLE     Plasmids with a kanamycin-resistance gene for site-directed 
            mutagenesis using the oligodeoxyribonucleotide-directed dual amber 
            method.
  JOURNAL   Gene 1995;167:333-4.
  PUBMED    8566803
REFERENCE   2  (bases 1 to 2204)
  AUTHORS   TaKaRa
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 2204)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Gene"; 
            date: "1995"; volume: "167"; pages: "333-4"
COMMENT     SGRef: number: 2; type: "Journal Article"
COMMENT     An NdeI site overlaps the lacZ-alpha start codon.
FEATURES             Location/Qualifiers
     source          1..2204
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    18..74
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(75..91)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(310..1125)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=aph(3')-Ia
                     /note="KanR (with amber stop codons)"
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSKPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLA*A*SRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     rep_origin      1450..2038
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    2095..2116
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        2131..2161
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2169..2185
                     /label=lac operator
                     /note="The lac repressor binds to the lac operator to
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
ORIGIN
        1 atgaccatga ttacgccgaa ttcgagctcg gtacccgggg atcctctaga gtcgacctgc
       61 aggcatgcaa gcttactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt
      121 acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag
      181 gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg cgatttattc
      241 aacaaagccg ccgtcccgtc aagtcagcgt aatgctctgc cagtgttaca accaattaac
      301 caattctgat tagaaaaact catcgagcat caaatgaaac tgcaatttat tcatatcagg
      361 attatcaata ccatattttt gaaaaagccg tttctgtaat gaaggagaaa actcaccgag
      421 gcagttccat aggatggcaa gatcctggta tcggtctgcg attccgactc gtccaacatc
      481 aatacaacct attaatttcc cctcgtcaaa aataaggtta tcaagtgaga aatcaccatg
      541 agtgacgact gaatccggtg agaatggcaa gagcttatgc atttctttcc agacttgttc
      601 aacaggccag ccattacgct cgtcatcaaa atcactcgca tcaaccaaac cgttattcat
      661 tcgtgactac gcctaagcga gacgaaatac gcgatcgctg ttaaaaggac aattacaaac
      721 aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca tcaacaatat tttcacctga
      781 atcaggatat tcttctaata cctggaatgc tgtttttccg gggatcgcag tggtgagtaa
      841 ccatgcatca tcaggagtac ggataaaatg cttgatggtc ggaagaggca taaattccgt
      901 cagccagttt agtctgacca tctcatctgt aacatcattg gcaacgctac ctttgccatg
      961 tttcagaaac aactctggcg catcgggctt cccatacaac cgatagattg tcgcacctga
     1021 ttgcccgaca ttatcgcgag cccatttata cccatataaa tcagcatcca tgttggaatt
     1081 taatcgcggc ttcgagcaag acgtttcccg ttgaatatgg ctcataacac cccttgtatt
     1141 actgtttatg taagcagaca gttttattgt tcatgatgat atatttttat cttgtgcaat
     1201 gtaacatcag agattttgag acacaacgtg gctttgttga ataaatcgaa cttttgctga
     1261 gttgaaggat cagatcacgc atcttcccga caacgcagac cgttccgtgc gtggctccct
     1321 cactttctgg ctggatgatg gggcgattca ggcctggtat gagtcagcaa caccttcttc
     1381 acgaggcaga cctctcgacg gatcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     1441 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     1501 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     1561 aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg
     1621 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     1681 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     1741 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     1801 gcgaacgacc tacaccgaac tgagatacct acagcgtgag cattgagaaa gcgccacgct
     1861 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     1921 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     1981 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     2041 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcataatgt
     2101 gagttagctc actcattagg caccccaggc tttacacttt atgcttccgg ctcgtatgtt
     2161 gtgtggaatt gtgagcggat aacaatttca cacaggaaac acat
//
