LOCUS       sequence-176906-        7660 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses lambda-red system with hyg selection marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7660)
  AUTHORS   Huang TW, Lam I, Chang HY, Tsai SF, Palsson BO, Charusanti P
  TITLE     Capsule deletion via a lambda-Red knockout system perturbs biofilm 
            formation and fimbriae expression in Klebsiella pneumoniae MGH 
            78578.
  JOURNAL   BMC Res Notes. 2014 Jan 8;7:13. doi: 10.1186/1756-0500-7-13.
  PUBMED    24398052
REFERENCE   2  (bases 1 to 7660)
  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: "BMC Res 
            Notes."; date: "2014-01-8"; volume: "7:13. doi"; pages: " 
            10.1186/1756-0500-7-13"
COMMENT     Sequence Label: pACBSR-hyg
FEATURES             Location/Qualifiers
     source          1..7660
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(461..745)
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     complement(519..538)
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             772..1650
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     primer_bind     complement(1830..1849)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     complement(1987..2009)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2150..2169
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     CDS             complement(2296..3321)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             complement(2296..3321)
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     promoter        complement(3322..3426)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3729..3746)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3863..4408)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(3943..3962)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        complement(4658..4749)
                     /gene="bla"
                     /label=AmpR promoter
     terminator      complement(4768..4795)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(4887..4973)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     5126..5143
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      complement(5203..5447)
                     /label=lambda tL3 terminator
                     /note="transcription terminator tL3 from phage lambda"
     CDS             complement(5448..6128)
                     /codon_start=1
                     /gene="exo"
                     /product="5' to 3' double-stranded DNA exonuclease in the 
                     lambda Red system"
                     /label=Exo
                     /note="creates 3' single-stranded DNA tails"
                     /translation="MTPDIILQRTGIDVRAVEQGDDAWHKLRLGVITASEVHNVIAKPR
                     SGKKWPDMKMSYFHTLLAEVCTGVAPEVNAKALAWGKQYENDARTLFEFTSGVNVTESP
                     IIYRDESMRTACSPDGLCSDGNGLELKCPFTSRDFMKFRLGGFEAIKSAYMAQVQYSMW
                     VTRKNAWYFANYDPRMKREGLHYVVIERDEKYMASFDEIVPEFIEKMDEALAEIGFVFG
                     EQWR"
     CDS             complement(6125..6910)
                     /codon_start=1
                     /gene="bet"
                     /product="single-stranded DNA binding recombinase in the 
                     lambda Red system"
                     /label=Beta
                     /note="mediates pairing of a single-stranded DNA with a 
                     complementary target"
                     /translation="MSTALATLAGKLAERVGMDSVDPQELITTLRQTAFKGDASDAQFI
                     ALLIVANQYGLNPWTKEIYAFPDKQNGIVPVVGVDGWSRIINENQQFDGMDFEQDNESC
                     TCRIYRKDRNHPICVTEWMDECRREPFKTREGREITGPWQSHPKRMLRHKAMIQCARLA
                     FGFAGIYDKDEAERIVENTAYTAERQPERDITPVNDETMQEINTLLIALDKTWDDDLLP
                     LCSQIFRRDIRASSELTQAEAVKALGFLKQKAAEQKVAA"
     CDS             complement(6916..7332)
                     /codon_start=1
                     /gene="gam"
                     /product="inhibitor of the host RecBCD nuclease in the 
                     lambda Red system"
                     /label=Gam
                     /note="protects the double-stranded DNA substrate for 
                     recombination"
                     /translation="MDINTETEIKQKHSLTPFPVFLISPAFRGRYFHSYFRSSAMNAYY
                     IQDRLEAQSWARHYQQLAREEKEAELADDMEKGLPQHLFESLCIDHLQRHGASKKSITR
                     AFDDDVEFQERMAEHIRYMVETIAHHQVDIDSEV"
     RBS             7341..7349
                     /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)"
ORIGIN
        1 gtatgccaga gacatcgggg tcaggtagtt ttcaaccagg ttgttcggga tggttttttt
       61 gttgttaacg atgaacaggt tagccagttt gttgaaagct tggtgtttga aagtctgggc
      121 gccccaggtg attaccaggt tacccaggtg gttaacacgt tcttttttgt gcggcgggga
      181 cagtacccac tgatcgtaca gcagacatac gtggtccatg tatgctttgt ttttccactc
      241 gaactgcata cagtaggttt taccttcatc acgagaacgg atgtaagcat cacccaggat
      301 cagaccgata cctgcttcga actgttcgat gttcagttcg atcagctggg atttgtattc
      361 tttcagcagt ttagagttcg gacccaggtt cattacctgg ttttttttga tgtttttcat
      421 atgcatatag gtaccctcct gctagcccaa aaaaacgggt atggagaaac agtagagagt
      481 tgcgataaaa agcgtcaggt aggatccgct aatcttatgg ataaaaatgc tatggcatag
      541 caaagtgtga cgccgtgcaa ataatcaatg tggacttttc tgccgtgatt atagacactt
      601 ttgttacgcg tttttgtcat ggctttggtc ccgctttgtt acagaatgct tttaataagc
      661 ggggttaccg gtttggttag cgagaagagc cagtaaaaga cgcagtgacg gcaatgtctg
      721 atgcaatatg gacaattggt ttcttctctg aatggcggga gtatgaaaag tatggctgaa
      781 gcgcaaaatg atcccctgct gccgggatac tcgtttaatg cccatctggt ggcgggttta
      841 acgccgattg aggccaacgg ttatctcgat ttttttatcg accgaccgct gggaatgaaa
      901 ggttatattc tcaatctcac cattcgcggt cagggggtgg tgaaaaatca gggacgagaa
      961 tttgtttgcc gaccgggtga tattttgctg ttcccgccag gagagattca tcactacggt
     1021 cgtcatccgg aggctcgcga atggtatcac cagtgggttt actttcgtcc gcgcgcctac
     1081 tggcatgaat ggcttaactg gccgtcaata tttgccaata cggggttctt tcgcccggat
     1141 gaagcgcacc agccgcattt cagcgacctg tttgggcaaa tcattaacgc cgggcaaggg
     1201 gaagggcgct attcggagct gctggcgata aatctgcttg agcaattgtt actgcggcgc
     1261 atggaagcga ttaacgagtc gctccatcca ccgatggata atcgggtacg cgaggcttgt
     1321 cagtacatca gcgatcacct ggcagacagc aattttgata tcgccagcgt cgcacagcat
     1381 gtttgcttgt cgccgtcgcg tctgtcacat cttttccgcc agcagttagg gattagcgtc
     1441 ttaagctggc gcgaggacca acgtatcagc caggcgaagc tgcttttgag caccacccgg
     1501 atgcctatcg ccaccgtcgg tcgcaatgtt ggttttgacg atcaactcta tttctcgcgg
     1561 gtatttaaaa aatgcaccgg ggccagcccg agcgagttcc gtgccggttg tgaagaaaaa
     1621 gtgaatgatg tagccgtcaa gttgtcataa ttggtaacga atcagacaat tgacggcttg
     1681 acggagtagc atagggtttg cagaatccct gcttcgtcca tttgacaggc acattatgca
     1741 tcgatgataa gctgtcaaac atgagcagat cctctacgcc ggacgcatcg tggccggcat
     1801 caccggcgcc acaggtgcgg ttgctggcgc ctatatcgcc gacatcaccg atggggaaga
     1861 tcgggctcgc cacttcgggc tcatgagcgc ttgtttcggc gtgggtatgg tggcaggccc
     1921 cgtggccggg ggactgttgg gcgccatctc cttctgcctc gcgcgtttcg gtgatgacgg
     1981 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
     2041 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggcgcagc
     2101 catgacccag tcacgtagcg atagcggagt gtatactggc ttaactatgc ggcatcagag
     2161 cagattgtac tgagagtgca cctgccattc atccgcttat tatcacttat tcaggcgtag
     2221 caaccaggcg tttaagggca ccaataactg ccttaaaaaa actcgaggta tatatgagta
     2281 aacttggtct gacagctatt cctttgccct cggacgagtg ctggggcgtc ggtttccact
     2341 atcggcgagt acttctacac agccatcggt ccagacggcc gcgcttctgc gggcgatttg
     2401 tgtacgcccg acagtcccgg ctccggatcg gacgattgcg tcgcatcgac cctgcgccca
     2461 agctgcatca tcgaaattgc cgtcaaccaa gctctgatag agttggtcaa gaccaatgcg
     2521 gagcatatac gcccggagcc gcggcgatcc tgcaagctcc ggatgcctcc gctcgaagta
     2581 gcgcgtctgc tgctccatac aagccaacca cggcctccag aagaagatgt tggcgacctc
     2641 gtattgggaa tccccgaaca tcgcctcgct ccagtcaatg accgctgtta tgcggccatt
     2701 gtccgtcagg acattgttgg agccgaaatc cgcgtgcacg aggtgccgga cttcggggca
     2761 gtcctcggcc caaagcatca gctcatcgag agcctgcgcg acggacgcac tgacggtgtc
     2821 gtccatcaca gtttgccagt gatacacatg gggatcagca atcgcgcata tgaaatcacg
     2881 ccatgtagtg tattgaccga ttccttgcgg tccgaatggg ccgaacccgc tcgtctggct
     2941 aagatcggcc gcagcgatcg catccatggc ctccgcgacc ggctgcagaa cagcgggcag
     3001 ttcggtttca ggcaggtctt gcaacgtgac accctgtgca cggcgggaga tgcaataggt
     3061 caggctctcg ctgaattccc caatgtcaag cacttccgga atcgggagcg cggccgatgc
     3121 aaagtgccga taaacataac gatctttgta gaaaccatcg gcgcagctat ttacccgcag
     3181 gacatatcca cgccctccta catcgaagct gaaagcacga gattcttcgc cctccgagag
     3241 ctgcatcagg tcggagacgc tgtcgaactt ttcgatcaga aacttctcga cagacgtcgc
     3301 ggtgagttca ggctttttca tactcttcct ttttcaatat tattgaagca tttatcaggg
     3361 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     3421 tccgcgcaca tttccccgag atctgcaaag tgcgtcgggt gatgctgcca acttactgat
     3481 ttagtgtatg atggtgtttt tgaggtgctc cagtggcttc tgtttctatc agctgtccct
     3541 cctgttcagc tactgacggg gtggtgcgta acggcaaaag caccgccgga catcagcgct
     3601 agcggagtgt atactggctt actatgttgg cactgatgag ggtgtcagtg aagtgcttca
     3661 tgtggcagga gaaaaaaggc tgcaccggtg cgtcagcaga atatgtgata caggatatat
     3721 tccgcttcct cgctcactga ctcgctacgc tcggtcgttc gactgcggcg agcggaaatg
     3781 gcttacgaac ggggcggaga tttcctggaa gatgccagga agatacttaa cagggaagtg
     3841 agagggccgc ggcaaagccg tttttccata ggctccgccc ccctgacaag catcacgaaa
     3901 tctgacgctc aaatcagtgg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     3961 cccctggcgg ctccctcgtg cgctctcctg ttcctgcctt tcggtttacc ggtgtcattc
     4021 cgctgttatg gccgcgtttg tctcattcca cgcctgacac tcagttccgg gtaggcagtt
     4081 cgctccaagc tggactgtat gcacgaaccc cccgttcagt ccgaccgctg cgccttatcc
     4141 ggtaactatc gtcttgagtc caacccggaa agacatgcaa aagcaccact ggcagcagcc
     4201 actggtaatt gatttagagg agttagtctt gaagtcatgc gccggttaag gctaaactga
     4261 aaggacaagt tttggtgact gcgctcctcc aagccagtta cctcggttca aagagttggt
     4321 agctcagaga accttcgaaa aaccgccctg caaggcggtt ttttcgtttt cagagcaaga
     4381 gattacgcgc agaccaaaac gatctcaaga agatcatctt attaatcaga taaaatattt
     4441 ctagatttca gtgcaattta tctcttcaaa tgtagcacct gaagtcagcc ccatacgata
     4501 taagttgtaa ttctcatgtt tgacagctta tcatcggtgc acccaactga tcttcagcat
     4561 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     4621 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     4681 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     4741 ataaacaaaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta
     4801 atttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc
     4861 gcaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa
     4921 caacagataa aacgaaaggc ccagtctttc gactgagcct ttcgttttat ttgatgcctg
     4981 gcagttccct actctcgcat ggggagaccc cacactacca tcggcgctac ggcgtttcac
     5041 ttctgagttc ggcatggggt caggtgggac caccgcgcta ctgccgccag gcaaattctg
     5101 ttttatcaga ccgcttctgc gttctgattt aatctgtatc aggctgaaaa tcttctctca
     5161 tccgccaaaa cagccaagct tgcatgcatt cttcgtctgt ttctactggt attggcacaa
     5221 acctgattcc aatttgagca aggctatgtg ccatctcgat actcgttctt aactcaacag
     5281 aagatgcttt gtgcatacag cccctcgttt attatttatc tcctcagcca gccgctgtgc
     5341 tttcagtgga tttcggataa cagaaaggcc gggaaatacc cagcctcgct ttgtaacgga
     5401 gtagacgaaa gtgattgcgc ctacccggat attatcgtga ggatgcgtca tcgccattgc
     5461 tccccaaata caaaaccaat ttcagccagt gcctcgtcca ttttttcgat gaactccggc
     5521 acgatctcgt caaaactcgc catgtacttt tcatcccgct caatcacgac ataatgcagg
     5581 ccttcacgct tcatacgcgg gtcatagttg gcaaagtacc aggcattttt tcgcgtcacc
     5641 cacatgctgt actgcacctg ggccatgtaa gctgacttta tggcctcgaa accaccgagc
     5701 cggaacttca tgaaatcccg ggaggtaaac gggcatttca gttcaaggcc gttgccgtca
     5761 ctgcataaac catcgggaga gcaggcggta cgcatacttt cgtcgcgata gatgatcggg
     5821 gattcagtaa cattcacgcc ggaagtgaat tcaaacaggg ttctggcgtc gttctcgtac
     5881 tgttttcccc aggccagtgc tttagcgtta acttccggag ccacaccggt gcaaacctca
     5941 gcaagcaggg tgtggaagta ggacattttc atgtcaggcc acttctttcc ggagcggggt
     6001 tttgctatca cgttgtgaac ttctgaagcg gtgatgacgc cgagccgtaa tttgtgccac
     6061 gcatcatccc cctgttcgac agctctcaca tcgatcccgg tacgctgcag gataatgtcc
     6121 ggtgtcatgc tgccaccttc tgctctgcgg ctttctgttt caggaatcca agagctttta
     6181 ctgcttcggc ctgtgtcagt tctgacgatg cacgaatgtc gcggcgaaat atctgggaac
     6241 agagcggcaa taagtcgtca tcccatgttt tatccagggc gatcagcaga gtgttaatct
     6301 cctgcatggt ttcatcgtta accggagtga tgtcgcgttc cggctgacgt tctgcagtgt
     6361 atgcagtatt ttcgacaatg cgctcggctt catccttgtc atagatacca gcaaatccga
     6421 aggccagacg ggcacactga atcatggctt tatgacgtaa catccgtttg ggatgcgact
     6481 gccacggccc cgtgatttct ctgccttcgc gagttttgaa tggttcgcgg cggcattcat
     6541 ccatccattc ggtaacgcag atcggatgat tacggtcctt gcggtaaatc cggcatgtac
     6601 aggattcatt gtcctgctca aagtccatgc catcaaactg ctggttttca ttgatgatgc
     6661 gggaccagcc atcaacgccc accaccggaa cgatgccatt ctgcttatca ggaaaggcgt
     6721 aaatttcttt cgtccacgga ttaaggccgt actggttggc aacgatcagt aatgcgatga
     6781 actgcgcatc gctggcatca cctttaaatg ccgtctggcg aagagtggtg atcagttcct
     6841 gtgggtcgac agaatccatg ccgacacgtt cagccagctt cccagccagc gttgcgagtg
     6901 cagtactcat tcgttttata cctctgaatc aatatcaacc tggtggtgag caatggtttc
     6961 aaccatgtac cggatgtgtt ctgccatgcg ctcctgaaac tcaacatcgt catcaaacgc
     7021 acgggtaatg gattttttgc tggccccgtg gcgttgcaaa tgatcgatgc atagcgattc
     7081 aaacaggtgc tggggcaggc ctttttccat gtcgtctgcc agttctgcct ctttctcttc
     7141 acgggcgagc tgctggtagt gacgcgccca gctctgagcc tcaagacgat cctgaatgta
     7201 ataagcgttc atggctgaac tcctgaaata gctgtgaaaa tatcgcccgc gaaatgccgg
     7261 gctgattagg aaaacaggaa agggggttag tgaatgcttt tgcttgatct cagtttcagt
     7321 attaatatcc attttttata acctccttag catgcctgca gacgtcgggc ccttatttca
     7381 ggaaagtttc ggaggagata gtgttcggca gtttgtacat catctgcggg atcaggtacg
     7441 gtttgatcag gttgtagaag atcaggtaag acatagaatc gatgtagatg atcggtttgt
     7501 ttttgttgat ttttacgtaa cagttcagtt ggaatttgtt acgcagaccc ttaaccaggt
     7561 attctacttc ttcgaaagtg aaagactggg tgttcagtac gatcgatttg ttggtagagt
     7621 ttttgttgta atcccattta ccaccatcat ccatgaacca
//
