LOCUS       sequence-98427-e        5238 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid that allows an arabinose-inducible expression of the I-SceI 
            homing endonuclease - for the en passant (two-step Red-mediated) 
            mutagenesis .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5238)
  AUTHORS   Tischer BK, von Einem J, Kaufer B, Osterrieder N
  TITLE     Two-step red-mediated recombination for versatile high-efficiency 
            markerless DNA manipulation in Escherichia coli.
  JOURNAL   Biotechniques. 2006 Feb;40(2):191-7.
  PUBMED    16526409
REFERENCE   2  (bases 1 to 5238)
  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: 
            "Biotechniques."; date: "2006-02"; volume: "40(2)"; pages: "191-7"
COMMENT     Sequence Label: pBAD-I-SceI
FEATURES             Location/Qualifiers
     source          1..5238
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(46..63)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      216..302
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      394..421
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        440..531
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(532..600,601..1392)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(750..769)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1434..1889
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1521..1540)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      2000..2588
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2489..2508
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2742..2759
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     2925..2947
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3086..3105
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     CDS             complement(3285..4163)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        4190..4474
                     /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     4397..4416
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
ORIGIN
        1 ggcatgcaag cttggctgtt ttggcggatg agagaagatt ttcagcctga tacagattaa
       61 atcagaacgc agaagcggtc tgataaaaca gaatttgcct ggcggcagta gcgcggtggt
      121 cccacctgac cccatgccga actcagaagt gaaacgccgt agcgccgatg gtagtgtggg
      181 gtctccccat gcgagagtag ggaactgcca ggcatcaaat aaaacgaaag gctcagtcga
      241 aagactgggc ctttcgtttt atctgttgtt tgtcggtgaa cgctctcctg agtaggacaa
      301 atccgccggg agcggatttg aacgttgcga agcaacggcc cggagggtgg cgggcaggac
      361 gcccgccata aactgccagg catcaaatta agcagaaggc catcctgacg gatggccttt
      421 ttgcgtttct acaaactctt ttgtttattt ttctaaatac attcaaatat gtatccgctc
      481 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
      541 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
      601 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
      661 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
      721 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtgttgac
      781 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
      841 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
      901 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
      961 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     1021 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     1081 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     1141 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     1201 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     1261 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     1321 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     1381 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttacgcgcc
     1441 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     1501 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     1561 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     1621 acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc
     1681 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     1741 gttccaaact tgaacaacac tcaaccctat ctcgggctat tcttttgatt tataagggat
     1801 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     1861 ttttaacaaa atattaacgt ttacaattta aaaggatcta ggtgaagatc ctttttgata
     1921 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     1981 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     2041 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     2101 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc
     2161 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     2221 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     2281 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     2341 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     2401 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     2461 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     2521 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     2581 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     2641 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     2701 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
     2761 aagcggaaga gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
     2821 gcatagggtc atggctgcgc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
     2881 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
     2941 tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag caaggagatg gcgcccaaca
     3001 gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga
     3061 agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac
     3121 ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatctgc tcatgtttga
     3181 cagcttatca tcgatgcata atgtgcctgt caaatggacg aagcagggat tctgcaaacc
     3241 ctatgctact ccgtcaagcc gtcaattgtc tgattcgtta ccaattatga caacttgacg
     3301 gctacatcat tcactttttc ttcacaaccg gcacggaact cgctcgggct ggccccggtg
     3361 cattttttaa atacccgcga gaaatagagt tgatcgtcaa aaccaacatt gcgaccgacg
     3421 gtggcgatag gcatccgggt ggtgctcaaa agcagcttcg cctggctgat acgttggtcc
     3481 tcgcgccagc ttaagacgct aatccctaac tgctggcgga aaagatgtga cagacgcgac
     3541 ggcgacaagc aaacatgctg tgcgacgctg gcgatatcaa aattgctgtc tgccaggtga
     3601 tcgctgatgt actgacaagc ctcgcgtacc cgattatcca tcggtggatg gagcgactcg
     3661 ttaatcgctt ccatgcgccg cagtaacaat tgctcaagca gatttatcgc cagcagctcc
     3721 gaatagcgcc cttccccttg cccggcgtta atgatttgcc caaacaggtc gctgaaatgc
     3781 ggctggtgcg cttcatccgg gcgaaagaac cccgtattgg caaatattga cggccagtta
     3841 agccattcat gccagtaggc gcgcggacga aagtaaaccc actggtgata ccattcgcga
     3901 gcctccggat gacgaccgta gtgatgaatc tctcctggcg ggaacagcaa aatatcaccc
     3961 ggtcggcaaa caaattctcg tccctgattt ttcaccaccc cctgaccgcg aatggtgaga
     4021 ttgagaatat aacctttcat tcccagcggt cggtcgataa aaaaatcgag ataaccgttg
     4081 gcctcaatcg gcgttaaacc cgccaccaga tgggcattaa acgagtatcc cggcagcagg
     4141 ggatcatttt gcgcttcagc catacttttc atactcccgc cattcagaga agaaaccaat
     4201 tgtccatatt gcatcagaca ttgccgtcac tgcgtctttt actggctctt ctcgctaacc
     4261 aaaccggtaa ccccgcttat taaaagcatt ctgtaacaaa gcgggaccaa agccatgaca
     4321 aaaacgcgta acaaaagtgt ctataatcac ggcagaaaag tccacattga ttatttgcac
     4381 ggcgtcacac tttgctatgc catagcattt ttatccataa gattagcgga tcctacctga
     4441 cgctttttat cgcaactctc tactgtttct ccatacccgt ttttttgggc tagcaggagg
     4501 gtacctatat gcatatgaaa aacatcaaaa aaaaccaggt aatgaacctg ggtccgaact
     4561 ctaaactgct gaaagaatac aaatcccagc tgatcgaact gaacatcgaa cagttcgaag
     4621 caggtatcgg tctgatcctg ggtgatgctt acatccgttc tcgtgatgaa ggtaaaacct
     4681 actgtatgca gttcgagtgg aaaaacaaag catacatgga ccacgtatgt ctgctgtacg
     4741 atcagtgggt actgtccccg ccgcacaaaa aagaacgtgt taaccacctg ggtaacctgg
     4801 taatcacctg gggcgcccag actttcaaac accaagcttt caacaaactg gctaacctgt
     4861 tcatcgttaa caacaaaaaa accatcccga acaacctggt tgaaaactac ctgaccccga
     4921 tgtctctggc atactggttc atggatgatg gtggtaaatg ggattacaac aaaaactcta
     4981 ccaacaaatc gatcgtactg aacacccagt ctttcacttt cgaagaagta gaatacctgg
     5041 ttaagggtct gcgtaacaaa ttccaactga actgttacgt aaaaatcaac aaaaacaaac
     5101 cgatcatcta catcgattct atgtcttacc tgatcttcta caacctgatc aaaccgtacc
     5161 tgatcccgca gatgatgtac aaactgccga acactatctc ctccgaaact ttcctgaaat
     5221 aagggcccga cgtctgca
//
