LOCUS       sequence-245914-        7744 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Vector for gene expression in Xylella fastidiosa, includes pemI/pemK
            toxin-antitoxin system for plasmid stability, Gateway cloning 
            compatible.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7744)
  AUTHORS   Burbank LP, Stenger DC
  TITLE     Plasmid Vectors for Xylella fastidiosa Utilizing a Toxin-Antitoxin 
            System for Stability in the Absence of Antibiotic Selection.
  JOURNAL   Phytopathology. 2016 Aug;106(8):928-36. doi: 
            10.1094/PHYTO-02-16-0097-R. Epub 2016 Jun 7.
  PUBMED    27088393
REFERENCE   2  (bases 1 to 7744)
  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"; doi: 
            "10.1094/PHYTO-02-16-0097-R"; journalName: "Phytopathology"; date: 
            "2016-08"; volume: "106"; issue: "8"; pages: "928-36"
COMMENT     Sequence Label: pXf20pemIK-GW
FEATURES             Location/Qualifiers
     source          1..7744
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      17..605
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     506..525
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     759..776
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    893..914
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(929..934,935..952,953..959)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    967..983
                     /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     972..994
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     991..1007
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     991..1007
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             2435..2446
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     protein_bind    2484..2608
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(2633..2638,2639..2656,2657..2663)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             2717..3376
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2783..2802)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             3718..4023
                     /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     3902..3921
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(4064..4188)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(4963..4982)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4964..4982)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4989..5006)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4989..5005)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4998..5020)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5146..5574
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5233..5252)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5443..5464
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             5918..6712
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5972..5991)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6582..6601
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     CDS             join(6730..6798,6799..7590)
                     /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(6948..6967)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
       61 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
      121 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
      181 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
      241 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
      301 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
      361 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
      421 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
      481 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
      541 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
      601 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
      661 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt
      721 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag
      781 cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
      841 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga
      901 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt
      961 gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca
     1021 agcttggtac cgagctcgga tccactagta acggccgcca gtgtgctgga attcggcttc
     1081 cataaagtta catgaaacgg attttttata aaaaatggtt tgtgaaagcg cagtgtgggc
     1141 aaaagcacgc ttaagcgtgc tggtgcacgt agtgcattgt ccatacggag cgttaagcgg
     1201 tggagcttgt gggcagagac ttgcggcggt gggcaaccgc ttgcggttgt ccacggacgt
     1261 gagtcggtgt gcgtagcgca ctgtccacaa atccacggct ttagttaaat ccgttgcttt
     1321 agattagaaa aagattaaag aagtattaga ggttatccac aggcaacgta agatattgat
     1381 tttatagtta aattttctta tttttcgtga aaattttagg atgctgcgac ggtaaaaagg
     1441 cgaatgctgc gacggtaaaa tctaggattc tgcgacggta catgaggatg ctgcgacggt
     1501 aaaaaggcga gtgctgcgac ggtaagttat ccacagattt caggggttaa tggcagcttg
     1561 gttcctacgt cacgggtaac gtatacaatt gacttaccac ctgggcggcg ctcaatcatg
     1621 tagctgaagt gctctccgtg gttcttacaa acgcttgcca gctcttcaag tgctggagct
     1681 aacgtctttt cttggaagtc cttcagagac ttagctgcgc tgccgcacag ctcatgaatc
     1741 ttggccacac tgtagggata gggttgacgg tgagttgcat agaatccatg aagccacttg
     1801 gccagagatg tatttagatc acatctaatc tgccaagaaa ccctcgtgta gccgtcgctt
     1861 tggtacaagg caacgatgcg ttgatttagt acaatgatgg cgcggcctgt ttcctcatca
     1921 tgataggctt cgtcgatcaa gctgccagag tacgtgtagc gtccttgtcg tacttcaagc
     1981 gcggttgctt gcagctcggc aagttgtcgt cgtagcactc gccgattgcc acctgtgtcc
     2041 ttcttgtcca agagtctcaa gagttgatag gtagtgactt cgcagcgatt gcctaactgc
     2101 tgttcgcgtg cgacgtgaac aaggcattcc cacacagtca aatagtcctg acataagcgg
     2161 attccggtgt acgtgatgga tgcgttttca acgctggcca tcacttggcg cttgaggtag
     2221 gggcgtttac cgcgttgaat cgcggcaaag agcgccgatc gaagtaagct gttcggaacg
     2281 ccgcgctgcg catctgtcca aattggcaat tgaagctgct gcaataggct cttttgcagg
     2341 ttggcttcct gtctggcacg cattttcgtt agtgaaagtt ctatgctgct cacgcttcct
     2401 ccttggttgt gtagtcacgc tcaagaacaa ctcgatagaa ggtcgagcgc gtgatgttgt
     2461 ataagccgaa ttctgcagat atcacaagtt tgtacaaaaa agctgaacga gaaacgtaaa
     2521 atgatataaa tatcaatata ttaaattaga ttttgcataa aaaacagact acataatact
     2581 gtaaaacaca acatatccag tcactatggc ggccgcatta ggcaccccag gctttacact
     2641 ttatgcttcc ggctcgtata atgtgtggat tttgagttag gatccgtcga gattttcagg
     2701 agctaaggaa gctaaaatgg agaaaaaaat cactggatat accaccgttg atatatccca
     2761 atggcatcgt aaagaacatt ttgaggcatt tcagtcagtt gctcaatgta cctataacca
     2821 gaccgttcag ctggatatta cggccttttt aaagaccgta aagaaaaata agcacaagtt
     2881 ttatccggcc tttattcaca ttcttgcccg cctgatgaat gctcatccgg aattccgtat
     2941 ggcaatgaaa gacggtgagc tggtgatatg ggatagtgtt cacccttgtt acaccgtttt
     3001 ccatgagcaa actgaaacgt tttcatcgct ctggagtgaa taccacgacg atttccggca
     3061 gtttctacac atatattcgc aagatgtggc gtgttacggt gaaaacctgg cctatttccc
     3121 taaagggttt attgagaata tgtttttcgt ctcagccaat ccctgggtga gtttcaccag
     3181 ttttgattta aacgtggcca atatggacaa cttcttcgcc cccgttttca ccatgggcaa
     3241 atattatacg caaggcgaca aggtgctgat gccgctggcg attcaggttc atcatgccgt
     3301 ttgtgatggc ttccatgtcg gcagaatgct taatgaatta caacagtact gcgatgagtg
     3361 gcagggcggg gcgtaaacgc gtggatccgg cttactaaaa gccagataac agtatgcgta
     3421 tttgcgcgct gatttttgcg gtataagaat atatactgat atgtataccc gaagtatgtc
     3481 aaaaagaggt atgctatgaa gcagcgtatt acagtgacag ttgacagcga cagctatcag
     3541 ttgctcaagg catatatgat gtcaatatct ccggtctggt aagcacaacc atgcagaatg
     3601 aagcccgtcg tctgcgtgcc gaacgctgga aagcggaaaa tcaggaaggg atggctgagg
     3661 tcgcccggtt tattgaaatg aacggctctt ttgctgacga gaacaggggc tggtgaaatg
     3721 cagtttaagg tttacaccta taaaagagag agccgttatc gtctgtttgt ggatgtacag
     3781 agtgatatta ttgacacgcc cgggcgacgg atggtgatcc ccctggccag tgcacgtctg
     3841 ctgtcagata aagtctcccg tgaactttac ccggtggtgc atatcgggga tgaaagctgg
     3901 cgcatgatga ccaccgatat ggccagtgtg ccggtctccg ttatcgggga agaagtggct
     3961 gatctcagcc accgcgaaaa tgacatcaaa aacgccatta acctgatgtt ctggggaata
     4021 taaatgtcag gctcccttat acacagccag tctgcaggtc gaccatagtg actggatatg
     4081 ttgtgtttta cagtattatg tagtctgttt tttatgcaaa atctaattta atatattgat
     4141 atttatatca ttttacgttt ctcgttcagc tttcttgtac aaagtggtga tatccatcac
     4201 actggcggcc gcgaattcac tagtgattgg ttgcagtggc ctgaacttgc cccaaaaatt
     4261 actggaagat ggtgacagct tttgccaaga catcggccaa gatgtttggt ggtaggcttt
     4321 ccactttgcg ggcattgcga gcgagtaagt cgagcgcacg tggttgatca catcgaacga
     4381 cgcccgttgt aatcgtacct agtaattcca cagcgaaccc gaggcgggag gcgaactcgc
     4441 cgccgtttgt gattggaagg atcaccggac aagaggtaag ccgattgaat cgctcactgg
     4501 agataacaac caccggacgg tgaccgcgtt gttcgtgtcc ggcggttggc tccagatcga
     4561 ccatgtaaac atcgccgcgt ttcatatcag ttcgcgtcct actgctggag catcgaccca
     4621 ttcacgttcc tcgggcggca acgggtaagc cccggaggct tccgcttcgg ccagcagttg
     4681 atccagcgtg tactgaggat gtaggactgg ctcaatgacc agacgaccgt ctgtgacggc
     4741 taagccgacc gttgcgccga cctcaagatg aagctgatca aggaaagcgg ggggaacggc
     4801 cagcatgatc gatccaccga ctttacgaag gtttgtggtg tgcatgatgg gcttctttcg
     4861 ttaagttata ttaaagtata atataattcc tccatttata cgcttatagc gaaaatcgaa
     4921 ttcccgcggc cgctcgagca tgcatctaga gggcccaatt cgccctatag tgagtcgtat
     4981 tacaattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa
     5041 cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc
     5101 accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggacgcg ccctgtagcg
     5161 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     5221 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     5281 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     5341 tcgaccccaa aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga
     5401 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa
     5461 ctggaacaac actcaaccct atctcggtct attcttttga tttataaggg attttgccga
     5521 tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca
     5581 aaattcaggg cgcaagggct gctaaaggaa gcggaacacg tagaaagcca gtccgcagaa
     5641 acggtgctga ccccggatga atgtcagcta ctgggctatc tggacaaggg aaaacgcaag
     5701 cgcaaagaga aagcaggtag cttgcagtgg gcttacatgg cgatagctag actgggcggt
     5761 tttatggaca gcaagcgaac cggaattgcc agctggggcg ccctctggta aggttgggaa
     5821 gccctgcaaa gtaaactgga tggctttctt gccgccaagg atctgatggc gcaggggatc
     5881 aagatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca
     5941 cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac
     6001 aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt
     6061 tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag cgcggctatc
     6121 gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg
     6181 aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat cccaccttgc
     6241 tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc
     6301 ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat
     6361 ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc
     6421 cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca
     6481 tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga
     6541 ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat
     6601 tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc
     6661 tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct gaattgaaaa
     6721 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     6781 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     6841 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     6901 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
     6961 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     7021 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     7081 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     7141 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     7201 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     7261 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     7321 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     7381 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     7441 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     7501 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     7561 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     7621 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     7681 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     7741 gaaa
//
