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ID   PLBU3      preliminary; circular DNA; SYN; 5100 BP.
XX
AC   L08929; VB0049;
XX
DT   04-JUN-1993 (Rel. 7, Created)
DT   01-JUL-1995 (Rel. 12, Last updated, Version 1)
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DE   E. coli plasmid vector pLBU3 - complete.
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KW   cloning vector.
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OS   Cloning vector
OC   Artificial sequences; Cloning vehicles.
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RN   [1]
RP   1-5100
RC   pLBU3
RA   Gilbert W.;
RT   "Obtained from VecBase 3.0";
RL   Unpublished (1991).
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RN   [2]
RC   pACYC214 from pACYC184
RC   pBU8 from pACYC214
RC   pBACH from pBU8
RC   pBU10 from pACYC184 & pBACH
RC   pLBU1 from pBU10 & phage fd terminator & pAD16/30, linker
RC   pLBU12a from pBU10 & pBR322
RC   pLBU3 from pLBU1 & pLBU12a
RC   pGBU207 from pLBU3 & linker & phage T5
RA   Gentz R., Langner A., Chang A.C., Cohen S.N., Bujard H.;
RT   "Cloning and analysis of strong promoters is made possible by
RT   the downstream placement of a RNA termination signal";
RL   Proc. Natl. Acad. Sci. U.S.A. 78:4936-4940(1981).
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RN   [3]
RC   from pDS2 & promoters
RA   Deuschle U., Kammerer W., Gentz R., Bujard H.;
RT   "Promoters of Escherichia coli: a hierarchy of in vivo strength
RT   indicates alternate structures";
RL   EMBO J. 5:2987-2994(1986).
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RN   [4]
RC   pAD16/30 from linker
RA   Schaller H.;
RT   ;
RL   Unpublished (1981).
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CC   These data and their annotation were supplied to GenBank by Will
CC   Gilbert under the auspices of the GenBank Currator Program.
CC
CC   Assembled from pBR322, GenBank(48)PL3, GenBank:EcoLac
CC   by F. Pfeiffer. The junction around position 412 is mere
CC   speculation!
CC   Assembly protocol:
CC   1- 224 is from PL3 (including overlap to EcoLac)
CC   225-439 is 'N' in PL3. The corresponding region of
CC   EcoLac was inserted as specified by the
CC   authors. Unfortunately, it is 22 bases shorter than
CC   claimed by the authors.
CC   217- 411 is from GenBank:EcoLac
CC   409- 411 is a filled-in HpaII site  (C'CGG ===> CCG)
CC   412- 413 is xx, because overlap between EcoLac and fd is not
CC   sufficiently specified in the publication. Two 'x'
CC   were added to hit the indicated stop-codon for the
CC   beta-galactosidase peptide.
CC   414- 418 is a filled-in SalI site. (G'TCGAC ===> GTCGA)
CC   The authors missed to publish the exact sequence of
CC   the SalI linker!
CC   418- 422 is a filled-in HindIII site, which has been restored
CC   by blunt-end ligation to the filled-in SalI site.
CC   417- 834 is from PL3. It contains the fd-part between the
CC   published junction sequences. Also it contains the
CC   part of the Tet region of pBR322.
CC   429- 761 is from phage fd
CC   771-5102 is from pBR322
CC   This vector is a promoter probe vector. The beta-galactosidase
CC   and - after a strong terminator from phage fd - the Tc gene can
CC   be activated. The vector itself does not confer Tc resistance.
CC
CC   It is proposed that the cloning of strong promoter signals
CC   from phage t5 or other sources might require the downstream
CC   placement of comparably strong termination signal as the one
CC   present in pLBU3.
CC
CC   The authors describe a 160 bp fragment of unknown origin that
CC   slipped into their vector pBU12a and was found useful because
CC   it provided stop-codons in all three reading frames.
CC   Bases 15- 65 are from pBR325 5336-5386 and
CC   bases 66-134 are from pBR325 4517-4439 (c).
CC   NM (pLBU3)
CC   CM (yes)
CC   NA (ds-DNA)
CC   TP (circular)
CC   ST ()
CC   TY (plasmid)
CC   SP ()
CC   HO (E.coli)
CC   CP ()
CC   FN (cloning)(promoter analysis)
CC   SE ()
CC   PA (pBR322)(PL3)(PFD)(EcoLac)
CC   BR ()
CC   OF ()
CC   OR ()
XX
FH   Key             Location/Qualifiers
FH
FT   misc_feature    0..0
FT                   /note="1. pACYC184, lacPO/lacZ/lacI
FT                   -> pACYC214
FT                   1. pACYC214 HincII-HincII 780bp, lacPO/lacZ/lacI
FT                   -> pBU8
FT                   1. pBU8 HincII-HpaII, lacPO/lacZalpha
FT                   BamHI linker 6bp ggatcc:SalI linker 6bp gtcgac
FT                   -> pBACH
FT                   1. pACYC184 HindIII 4245bp 1525..1525, tet gene
FT                   blunt end:blunt end
FT                   2. pBACH BamHI-SalI, lacPO/lacZalpha
FT                   blunt end:blunt end
FT                   -> pBU10
FT                   1. pBU10 HindIII, 3' to lacZ
FT                   2. pAD16/30 HindIII-BamHI 28bp, linker
FT                   3. phage fd Sau3A/MboI-Sau3A/MboI 332bp 1382..1714,
FT                   \ terminator [338bp]
FT                   \ MboI= 1382 1714 2221 5646
FT                   4. pAD16/30 BamHI-HindIII 28bp, linker
FT                   -> pLBU1
FT                   1. pBU10 HpaII-HindIII, lac operator/lacZalpha
FT                   DNA polymerase I:
FT                   2. pBR322 remove EcoRI-HindIII 31bp 4360..4361..30,
FT                   \ 4330bp
FT                   DNA polymerase I:
FT                   -> pLBU12a
FT                   1. pLBU1 HindIII-SalI, fd terminator
FT                   2. pLBU12a remove HindIII-SalI
FT                   -> pLBU3 5100bp [unique EcoRI]"
FT   misc_feature    1..224
FT                   /note="from pL3"
FT   misc_feature    15..65
FT                   /note="from pBR325"
FT   misc_feature    66..134
FT                   /note="from pBR325"
FT   CDS             163..411
FT                   /note="GEN E. coli beta-galactosidase gene (lacZ)"
FT   misc_feature    409..418
FT                   /note="undefined junction (CCGxxGTCGA)"
FT   misc_feature    418..835
FT                   /note="from pL3"
FT   misc_feature    complement(429..761)
FT                   /note="from bacteriophage fd"
FT   misc_feature    771..5102
FT                   /note="from pBR322"
FT   misc_feature    828..2018
FT                   /note="ANT E. coli tetracycline resistance gene (tet);
FT                   but no promoter"
FT   CDS             4040..4828
FT                   /note="ANT E. coli beta-lactamase gene (bla)
FT                   ampicillin resistance gene (apr/amp)"
XX
SQ   Sequence 5100 BP; 1175 A; 1381 C; 1310 G; 1234 T; 0 other;
     gaattcggtc gcttgttcag ggcagggtcg ttaaatagcc gcttatgtct attgctggtt
     taccgcggta gtgatcttat ttcattatgg tgaaagttgg aacctcttac gtgccgatca
     acgtctcatt ttcgccaaaa gttgacctag aaacttcccg aactcgtatg ttgtgtggaa
     ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac ggattcactg
     gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt
     gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct
     tcccaacagt tgcgcagcct gaatggcgaa tggcgctttg cctggtttcc ggtcgaagct
     tggccggatc taaagttttg tcgtctttcc agacgttagt aaatgaattt tctgtatgag
     gttttgctaa acaactttca acagtttcag cggagtgaga atagaaagga acaactaaag
     gaattgcgaa taataatttt ttcacgttga aaatctccaa aaaaaaaggc tccaaaagga
     gcctttaatt gtatcggttt atcagcttgc tttcgaggtg aatttcttaa acagcttgat
     accgatagtt gcgccgacaa tgacaacaac catcgcccac gcataaccga tatattcggt
     cgctgaggct tgcagggagt caaaggccgc ttttgcgggg atccggccaa gctttaatgc
     ggtagtttat cacagttaaa ttgctaacgc agtcaggcac cgtgtatgaa atctaacaat
     gcgctcatcg tcatcctcgg caccgtcacc ctggatgctg taggcatagg cttggttatg
     ccggtactgc cgggcctctt gcgggatatc gtccattccg acagcatcgc cagtcactat
     ggcgtgctgc tagcgctata tgcgttgatg caatttctat gcgcacccgt tctcggagca
     ctgtccgacc gctttggccg ccgcccagtc ctgctcgctt cgctacttgg agccactatc
     gactacgcga tcatggcgac cacacccgtc ctgtggatcc tctacgccgg acgcatcgtg
     gccggcatca ccggcgccac aggtgcggtt gctggcgcct atatcgccga catcaccgat
     ggggaagatc gggctcgcca cttcgggctc atgagcgctt gtttcggcgt gggtatggtg
     gcaggccccg tggccggggg actgttgggc gccatctcct tgcatgcacc attccttgcg
     gcggcggtgc tcaacggcct caacctacta ctgggctgct tcctaatgca ggagtcgcat
     aagggagagc gtcgaccgat gcccttgaga gccttcaacc cagtcagctc cttccggtgg
     gcgcggggca tgactatcgt cgccgcactt atgactgtct tctttatcat gcaactcgta
     ggacaggtgc cggcagcgct ctgggtcatt ttcggcgagg accgctttcg ctggagcgcg
     acgatgatcg gcctgtcgct tgcggtattc ggaatcttgc acgccctcgc tcaagccttc
     gtcactggtc ccgccaccaa acgtttcggc gagaagcagg ccattatcgc cggcatggcg
     gccgacgcgc tgggctacgt cttgctggcg ttcgcgacgc gaggctggat ggccttcccc
     attatgattc ttctcgcttc cggcggcatc gggatgcccg cgttgcaggc catgctgtcc
     aggcaggtag atgacgacca tcagggacag cttcaaggat cgctcgcggc tcttaccagc
     ctaacttcga tcactggacc gctgatcgtc acggcgattt atgccgcctc ggcgagcaca
     tggaacgggt tggcatggat tgtaggcgcc gccctatacc ttgtctgcct ccccgcgttg
     cgtcgcggtg catggagccg ggccacctcg acctgaatgg aagccggcgg cacctcgcta
     acggattcac cactccaaga attggagcca atcaattctt gcggagaact gtgaatgcgc
     aaaccaaccc ttggcagaac atatccatcg cgtccgccat ctccagcagc cgcacgcggc
     gcatctcggg cagcgttggg tcctggccac gggtgcgcat gatcgtgctc ctgtcgttga
     ggacccggct aggctggcgg ggttgcctta ctggttagca gaatgaatca ccgatacgcg
     agcgaacgtg aagcgactgc tgctgcaaaa cgtctgcgac ctgagcaaca acatgaatgg
     tcttcggttt ccgtgtttcg taaagtctgg aaacgcggaa gtcagcgccc tgcaccatta
     tgttccggat ctgcatcgca ggatgctgct ggctaccctg tggaacacct acatctgtat
     taacgaagcg ctggcattga ccctgagtga tttttctctg gtcccgccgc atccataccg
     ccagttgttt accctcacaa cgttccagta accgggcatg ttcatcatca gtaacccgta
     tcgtgagcat cctctctcgt ttcatcggta tcattacccc catgaacaga aattccccct
     tacacggagg catcaagtga ccaaacagga aaaaaccgcc cttaacatgg cccgctttat
     cagaagccag acattaacgc ttctggagaa actcaacgag ctggacgcgg atgaacaggc
     agacatctgt gaatcgcttc acgaccacgc tgatgagctt taccgcagct gcctcgcgcg
     tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg
     tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg
     gtgtcggggc gcagccatga cccagtcacg tagcgatagc ggagtgtata ctggcttaac
     tatgcggcat cagagcagat tgtactgaga gtgcaccata tgcggtgtga aataccgcac
     agatgcgtaa ggagaaaata ccgcatcagg cgctcttccg cttcctcgct cactgactcg
     ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca
     gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     aatagtttgc gcaacgttgt tgccattgct gcaggcatcg tggtgtcacg ctcgtcgttt
     ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     cggcgaccga gttgctcttg cccggcgtca acacgggata ataccgcgcc acatagcaga
     actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc
     attattatca tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtcttcaa
//