LOCUS       sequence-77333-e        6198 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Kanamycin and erethromycin resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6198)
  AUTHORS   Tyurin MV, Desai SG, Lynd LR
  TITLE     Electrotransformation of Clostridium thermocellum.
  JOURNAL   Appl Environ Microbiol. 2004 Feb;70(2):883-90.
  PUBMED    14766568
REFERENCE   2  (bases 1 to 6198)
  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: "Appl 
            Environ Microbiol."; date: "2004-02"; volume: "70(2)"; pages: 
            "883-90"
COMMENT     Sequence Label: pIKM1
FEATURES             Location/Qualifiers
     source          1..6198
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             2581..3375
                     /codon_start=1
                     /gene="aphA-3"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
                     TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
                     EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
                     PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
                     FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
     primer_bind     complement(3569..3586)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3569..3585)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3578..3600)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(3794..3813)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3913..3935
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3973..3991)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4059..4163
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4164..4232,4233..5024)
                     /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     complement(4382..4401)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5195..5783
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5684..5703
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5937..5954
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6071..6092
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6107..6112,6113..6130,6131..6137)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6145..6161
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /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)."
     primer_bind     6150..6172
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6169..6185
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6169..6185
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 agctttggct aacacacacg ccattccaac caatagtttt ctcggcataa agccatgctc
       61 tgacgcttaa atgcactaat gccttaaaaa aacattaaag tctaacacac tagacttatt
      121 tacttcgtaa ttaagtcgtt aaaccgtgtg ctctacgacc aaaagtataa aacctttaag
      181 aactttcttt tttcttgtaa aaaaagaaac tagataaatc tctcatatct tttattcaat
      241 aatcgcatca gattgcagta taaatttaac gatcactcat catgttcata tttatcagag
      301 ctcgtgctat aattatacta attttataag gaggaaaaaa taaagagggt tataatgaac
      361 gagaaaaata taaaacacag tcaaaacttt attacttcaa aacataatat agataaaata
      421 atgacaaata taagattaaa tgaacatgat aatatctttg aaatcggctc aggaaaaggg
      481 cattttaccc ttgaattagt acagaggtgt aatttcgtaa ctgccattga aatagaccat
      541 aaattatgca aaactacaga aaataaactt gttgatcacg ataatttcca agttttaaac
      601 aaggatatat tgcagtttaa atttcctaaa aaccaatcct ataaaatatt tggtaatata
      661 ccttataaca taagtacgga tataatacgc aaaattgttt ttgatagtat agctgatgag
      721 atttatttaa tcgtggaata cgggtttgct aaaagattat taaatacaaa acgctcattg
      781 gcattatttt taatggcaga agttgatatt tctatattaa gtatggttcc aagagaatat
      841 tttcatccta aacctaaagt gaatagctca cttatcagat taaatagaaa aaaatcaaga
      901 atatcacaca aagataaaca gaagtataat tatttcgtta tgaaatgggt taacaaagaa
      961 tacaagaaaa tatttacaaa aaatcaattt aacaattcct taaaacatgc aggaattgac
     1021 gatttaaaca atattagctt tgaacaattc ttatctcttt tcaatagcta taaattattt
     1081 aataagtaag ttaagggatg cataaactgc atcccttaac ttgtttttcg tgtacctatt
     1141 ttttgtgaat cgattatgtc ttttgcgcat tcacttcttt tctatataaa tatgagcgaa
     1201 gcgaataagc gtcggaaaag cagcaaaaag tttccttttt gctgttggag catgggggtt
     1261 cagggggtgc agtatctgac gtcaatgccg agcgaaagcg agccgaaggg tagcatttac
     1321 gttagataac cccctgatat gctccgacgc tttatataga aaagaagatt caactaggta
     1381 aaatcttaat ataggttgag atgataaggt ttataaggaa tttgtttgtt ctaatttttc
     1441 actcattttg ttctaatttc ttttaacaaa tgttcttttt tttttagaac agttatgata
     1501 tagttagaat agtttaaaat aaggagtgag aaaaagatga aagaaagata tggaacagtc
     1561 tataaaggct ctcagaggct catagacgaa gaaagtggag aagtcataga ggtagacaag
     1621 ttataccgta aacaaacgtc tggtaacttc gtaaaggcat atatagtgca attaataagt
     1681 atgttagata tgattggcgg aaaaaaactt aaaatcgtta actatatcct agataatgtc
     1741 cacttaagta acaatacaat gatagctaca acaagagaaa tagcaaaagc tacaggaaca
     1801 agtctacaaa cagtaataac aacacttaaa atcttagaag aaggaaatat tataaaaaga
     1861 aaaactggag tattaatgtt aaaccctgaa ctactaatga gaggcgacga ccaaaaacaa
     1921 aaatacctct tactcgaatt tgggaacttt gagcaagagg caaatgaaat agattgacct
     1981 cccaataaca ccacgtagtt attgggaggt caatctatga aatgcgatta agcttggctg
     2041 caggtcgata aacccagcga accatttgag gtgataggta agattatacc gaggtatgaa
     2101 aacgagaatt ggacctttac agaattactc tatgaagcgc catatttaaa aagctaccaa
     2161 gacgaagagg atgaagagga tgaggaggca gattgccttg aatatattga caatactgat
     2221 aagataatat atcttttata tagaagatat cgccgtatgt aaggatttca gggggcaagg
     2281 cataggcagc gcgcttatca atatatctat agaatgggca aagcataaaa acttgcatgg
     2341 actaatgctt gaaacccagg acaataacct tatagcttgt aaattctatc ataattgtgg
     2401 tttcaaaatc ggctccgtcg atactatgtt atacgccaac tttcaaaaca actttgaaaa
     2461 agctgttttc tggtatttaa ggttttagaa tgcaaggaac agtgaattgg agttcgtctt
     2521 gttataatta gcttcttggg gtatctttaa atactgtaga aaagaggaag gaaataataa
     2581 atggctaaaa tgagaatatc accggaattg aaaaaactga tcgaaaaata ccgctgcgta
     2641 aaagatacgg aaggaatgtc tcctgctaag gtatataagc tggtgggaga aaatgaaaac
     2701 ctatatttaa aaatgacgga cagccggtat aaagggacca cctatgatgt ggaacgggaa
     2761 aaggacatga tgctatggct ggaaggaaag ctgcctgttc caaaggtcct gcactttgaa
     2821 cggcatgatg gctggagcaa tctgctcatg agtgaggccg atggcgtcct ttgctcggaa
     2881 gagtatgaag atgaacaaag ccctgaaaag attatcgagc tgtatgcgga gtgcatcagg
     2941 ctctttcact ccatcgacat atcggattgt ccctatacga atagcttaga cagccgctta
     3001 gccgaattgg attacttact gaataacgat ctggccgatg tggattgcga aaactgggaa
     3061 gaagacactc catttaaaga tccgcgcgag ctgtatgatt ttttaaagac ggaaaagccc
     3121 gaagaggaac ttgtcttttc ccacggcgac ctgggagaca gcaacatctt tgtgaaagat
     3181 ggcaaagtaa gtggctttat tgatcttggg agaagcggca gggcggacaa gtggtatgac
     3241 attgccttct gcgtccggtc gatcagggag gatatcgggg aagaacagta tgtcgagcta
     3301 ttttttgact tactggggat caagcctgat tgggagaaaa taaaatatta tattttactg
     3361 gatgaattgt tttagtacct agatttagat gtctaaaaag ctttaactac aagcttttta
     3421 gacatctaat cttttctgaa gtacatccgc aactgtccat actctgatgt tttatatctt
     3481 ttctaaaagt tcgctagata ggggtcccga gcgcctacga ggaatttgta tcgactctag
     3541 aggatccccg ggtaccgagc tcgaattcac tggccgtcgt tttacaacgt cgtgactggg
     3601 aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc
     3661 gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg
     3721 aatggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatat
     3781 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc cgacacccgc
     3841 caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag
     3901 ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg
     3961 cgagacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg ataataatgg
     4021 tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     4081 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     4141 aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct
     4201 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     4261 atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta
     4321 agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc
     4381 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
     4441 tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg
     4501 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     4561 ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca
     4621 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     4681 acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa
     4741 ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg gaggcggata
     4801 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     4861 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc
     4921 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     4981 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt
     5041 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     5101 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     5161 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     5221 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     5281 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     5341 tccttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     5401 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     5461 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     5521 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     5581 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     5641 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     5701 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     5761 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     5821 tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc
     5881 gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg
     5941 agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt
     6001 ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc
     6061 gcaacgcaat taatgtgagt tagctcactc attaggcacc ccaggcttta cactttatgc
     6121 ttccggctcg tatgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct
     6181 atgaccatga ttacgcca
//
