p11-LacY-wtx1

LOCUS       p11-LacY-wtx1                6092 bp ds-DNA     circular SYN 06-AUG-2018
DEFINITION  The reporter plasmid p11-LacY-wtx1 encodes the toxin CcdB gene under
            the inducible BAD promoter and a single copy of the endonuclease 
            cleavage site which is readily extendable to multiple copies..
ACCESSION   .
VERSION     .
KEYWORDS    p11-LacY-wtx1
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6092)
  AUTHORS   Chen Z, Zhao H
  TITLE     A highly sensitive selection method for directed evolution of homing
            endonucleases.
  JOURNAL   Nucleic Acids Res. 2005 Oct 6;33(18):e154.
  PUBMED    16214805
REFERENCE   2  (bases 1 to 6092)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 6, 2018 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6092
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     CDS             complement(95..973)
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     promoter        1000..1284
                     /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     1207..1226
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             1316..1621
                     /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     1500..1519
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     primer_bind     complement(1716..1733)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1886..1972
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2064..2091
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        2110..2201
                     /gene="bla"
                     /label=AmpR promoter
     CDS             2202..3062
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(2420..2439)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3104..3559
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3191..3210)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      3670..4258
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4159..4178
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4412..4429
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4594..4615
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4630..4660
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4668..4684
                     /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     4673..4695
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4692..4708
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4692..4708
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4724..4741)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4724..4740)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 tcgatgcata atgtgcctgt caaatggacg aagcagggat tctgcaaacc ctatgctact
       61 ccgtcaagcc gtcaattgtc tgattcgtta ccaattatga caacttgacg gctacatcat
      121 tcactttttc ttcacaaccg gcacggaact cgctcgggct ggccccggtg cattttttaa
      181 atacccgcga gaaatagagt tgatcgtcaa aaccaacatt gcgaccgacg gtggcgatag
      241 gcatccgggt ggtgctcaaa agcagcttcg cctggctgat acgttggtcc tcgcgccagc
      301 ttaagacgct aatccctaac tgctggcgga aaagatgtga cagacgcgac ggcgacaagc
      361 aaacatgctg tgcgacgctg gcgatatcaa aattgctgtc tgccaggtga tcgctgatgt
      421 actgacaagc ctcgcgtacc cgattatcca tcggtggatg gagcgactcg ttaatcgctt
      481 ccatgcgccg cagtaacaat tgctcaagca gatttatcgc cagcagctcc gaatagcgcc
      541 cttccccttg cccggcgtta atgatttgcc caaacaggtc gctgaaatgc ggctggtgcg
      601 cttcatccgg gcgaaagaac cccgtattgg caaatattga cggccagtta agccattcat
      661 gccagtaggc gcgcggacga aagtaaaccc actggtgata ccattcgcga gcctccggat
      721 gacgaccgta gtgatgaatc tctcctggcg ggaacagcaa aatatcaccc ggtcggcaaa
      781 caaattctcg tccctgattt ttcaccaccc cctgaccgcg aatggtgaga ttgagaatat
      841 aacctttcat tcccagcggt cggtcgataa aaaaatcgag ataaccgttg gcctcaatcg
      901 gcgttaaacc cgccaccaga tgggcattaa acgagtatcc cggcagcagg ggatcatttt
      961 gcgcttcagc catacttttc atactcccgc cattcagaga agaaaccaat tgtccatatt
     1021 gcatcagaca ttgccgtcac tgcgtctttt actggctctt ctcgctaacc aaaccggtaa
     1081 ccccgcttat taaaagcatt ctgtaacaaa gcgggaccaa agccatgaca aaaacgcgta
     1141 acaaaagtgt ctataatcac ggcagaaaag tccacattga ttatttgcac ggcgtcacac
     1201 tttgctatgc catagcattt ttatccataa gattagcgga tcctacctga cgctttttat
     1261 cgcaactctc tactgtttct ccatacccgt ttttttgggc tagcgattga aaacgatgca
     1321 gtttaaggtt tacacctata aaagagagag ccgttatcgt ctgtttgtgg atgtacagag
     1381 tgatattatt gacacgcccg ggcgacggat ggtgatcccc ctggccagtg cacgtctgct
     1441 gtcagataaa gtctcccgtg aactttaccc ggtggtgcat atcggggatg aaagctggcg
     1501 catgatgacc accgatatgg ccagtgtgcc ggtctccgtt atcggggaag aagtggctga
     1561 tctcagccac cgcgaaaatg acatcaaaaa cgccattaac ctgatgtttt ggggaatata
     1621 atctagcatt acgctaggga taacagggta atatcacgct ctagacatac ggcatgcaag
     1681 cttggctgtt ttggcggatg agagaagatt ttcagcctga tacagattaa atcagaacgc
     1741 agaagcggtc tgataaaaca gaatttgcct ggcggcagta gcgcggtggt cccacctgac
     1801 cccatgccga actcagaagt gaaacgccgt agcgccgatg gtagtgtggg gtctccccat
     1861 gcgagagtag ggaactgcca ggcatcaaat aaaacgaaag gctcagtcga aagactgggc
     1921 ctttcgtttt atctgttgtt tgtcggtgaa cgctctcctg agtaggacaa atccgccggg
     1981 agcggatttg aacgttgcga agcaacggcc cggagggtgg cgggcaggac gcccgccata
     2041 aactgccagg catcaaatta agcagaaggc catcctgacg gatggccttt ttgcgtttct
     2101 acaaactctt ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     2161 aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc
     2221 gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa
     2281 cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac
     2341 tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga
     2401 tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtgttgac gccgggcaag
     2461 agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca
     2521 cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca
     2581 tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa
     2641 ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc
     2701 tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgcagca atggcaacaa
     2761 cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag
     2821 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct
     2881 ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac
     2941 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa
     3001 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt
     3061 aactgtcaga ccaagtttac tcatatatac tttagattga tttacgcgcc ctgtagcggc
     3121 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     3181 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     3241 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     3301 gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     3361 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     3421 tgaacaacac tcaaccctat ctcgggctat tcttttgatt tataagggat tttgccgatt
     3481 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     3541 atattaacgt ttacaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     3601 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     3661 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     3721 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     3781 aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg
     3841 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     3901 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     3961 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     4021 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     4081 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     4141 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     4201 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     4261 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtt
     4321 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga
     4381 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga
     4441 gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgtac
     4501 gatttaaata ggcctgactc actataggga gaccggaatt ccctggcacg acaggtttcc
     4561 cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca ctcattaggg
     4621 accccgggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg tgagcggata
     4681 acaatttcac acaggaaaca gctatgacca tgattacgga ttcactggcc gtcgttttac
     4741 aacgtcgtga ctggtaaaac ccgggcgtta cccaacttaa tcgccttgca gcacatcccc
     4801 ctttcgccag caggcgtaat aaggaaagga tccatgtact atttgaaaaa cacaaacttt
     4861 tggatgttcg gtttattctt tttcttttac ttttttatca tgggagccta cttcccgttt
     4921 ttcccgattt ggctacatga tatcaaccat atcagcaaaa gtgatacggg tattattttt
     4981 gccgctattt ctctgttctc gctattattc caaccgctgt ttggtctgct ttctgacaaa
     5041 ctcggtctac gcaaatacct gctgtggatt attaccggca tgttagtgat gtttgcgccg
     5101 ttctttattt ttatcttcgg gccactgctg cagtacaaca ttttagtagg atcgattgtt
     5161 ggtggtattt atctaggctt tagttttaac gccggtgcgc cagcagtaga ggcatttatt
     5221 gagaaagtca gccggcgcag taatttcgaa tttggtcgcg cgcggatgtt tggcagtgtt
     5281 ggctgggcgc tggttgcctc gattgtcggg atcatgttca ccattaataa tcagtttgtt
     5341 ttctggctgg gctctggcag ttgtctcatc ctcgccgttt tactcttttt cgccaaaacg
     5401 gacgcgccct cgagtgccac ggttgccaat gcggtaggtg ccaaccattc ggcatttagc
     5461 cttaagctgg cactggaact gttcagacag ccaaaactgt ggtttttgtc actgtatgtt
     5521 attggcgttt cctccaccta cgatgttttt gaccaacagt ttgctaattt ctttacttcg
     5581 ttctttgcta ccggtgaaca gggtacccgc gtatttggct acgtaacgac aatgggcgaa
     5641 ttacttaacg cctcgattat gttctttgcg ccactgatca ttaatcgcat cggtgggaag
     5701 aatgccctgc tgctggctgg cactattatg tctgtacgta ttattggctc atcgttcgcc
     5761 acctcagcgc tggaagtggt tattctgaaa acgctgcata tgtttgaagt accgttcctg
     5821 ctggtgggct cctttaaata tattactagt cagtttgaag tgcgtttttc agcgacgatt
     5881 tatctggtca gtttcagctt ctttaagcaa ctggcgatga tttttatgtc tgtactggcg
     5941 ggcaatatgt atgaaagcat aggtttccaa ggcgcttatc tggtgctggg tctggtggcg
     6001 ctgggcttca ccttaatttc cgtgttcacg cttagcggcc cgggcccgct ttccctgctg
     6061 cgtcgtcagg tgaatgaagt cgcttaaagg cc
//