LOCUS JF934954 6295 bp DNA circular SYN 24-JAN-2013
DEFINITION Expression vector pLoxGentrc, complete sequence.
ACCESSION JF934954
VERSION JF934954.1
KEYWORDS .
SOURCE Expression vector pLoxGentrc
ORGANISM Expression vector pLoxGentrc
other sequences; artificial sequences; vectors.
REFERENCE 1 (bases 1 to 1632; 3874 to 6295)
AUTHORS Palmeros,B., Wild,J., Szybalski,W., Le Borgne,S.,
Hernandez-Chavez,G., Gosset,G., Valle,F. and Bolivar,F.
TITLE A family of removable cassettes designed to obtain
antibiotic-resistance-free genomic modifications of Escherichia
coli and other bacteria
JOURNAL Gene 247 (1-2), 255-264 (2000)
PUBMED 10773465
REFERENCE 2 (bases 1 to 6295)
AUTHORS Sabido,A., Martinez,L.M., de Anda,R., Martinez,A., Bolivar,F. and
Gosset,G.
TITLE A novel plasmid vector designed for chromosomal gene integration
and expression: Use for developing a genetically stable Escherichia
coli melanin production strain
JOURNAL Plasmid 69 (1), 16-23 (2013)
PUBMED 22884755
REFERENCE 3 (bases 1 to 6295)
AUTHORS Sabido,A. and Gosset,G.
TITLE Direct Submission
JOURNAL Submitted (11-MAY-2011) Cell Engineering and Biocatalysis,
Biotechnology Institute, Av. Universidad 2001, Cuernavaca, Morelos
62210, Mexico
FEATURES Location/Qualifiers
source 1..6295
/organism="Expression vector pLoxGentrc"
/mol_type="other DNA"
/db_xref="taxon:1031331"
/lab_host="Escherichia coli"
/note="derivative of vector pLoxGen4; numbered such that
the first nucleotide is the last nucleotide of intergenic
region of M13 bacteriophage and the count increases
clockwise; carries ampicillin (bla) and gentamycin (aacC1)
resistance genes; two parallel loxP sites from pSL1180 are
flanking the aacC1 gene; contains all sequences for the
correct expression of a gene from plasmid pTrc99A; carries
a ColE1 and M13 bacteriophage replication origins; used
for the integration of genes into Escherichia coli"
misc_feature 1..474
/note="intergenic region of M13 bacteriophage; includes
all necessary elements for single strand replication in M1
bacteriophage DNA"
misc_feature 688..762
/note="Multiple Cloning Site (MCS) modified from plasmid
pLoxGen4; contains the following restriction sites: PmlI,
SfiI, BalI, SpeI, ScaI, XhoI, SalI, ClaI and BamHI;
HindIII site was eliminated"
misc_feature complement(763..796)
/note="loxP site; the 34 bp loxP sequence is the DNA
target site for Cre site-specific recombinase from P1
bacteriophage; the two loxP sequences are flanking the
resistance gene; therefore, antibiotic resistance gene can
be eliminated by Cre trans-action"
gene complement(840..1590)
/gene="aacC1"
CDS complement(840..1373)
/gene="aacC1"
/function="confers resistance to gentamycin"
/codon_start=1
/transl_table=11
/product="gentamycin acetyltransferase"
/protein_id="AEG75266.1"
/translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAAL
DLFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPRFEQP
RSEIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKL
GIREEVMHFDIDPSTAT"
regulatory complement(1557..1563)
/regulatory_class="minus_10_signal"
/gene="aacC1"
regulatory complement(1585..1590)
/regulatory_class="minus_35_signal"
/gene="aacC1"
misc_feature complement(1599..1632)
/note="loxP site; DNA target site for Cre site-specific
recombinase"
misc_feature complement(1697..2122)
/note="derived from the rrnB operon; the entire operon is
not contained in this vector"
gene complement(1716..2041)
/gene="rrnB"
regulatory complement(1716..1743)
/regulatory_class="terminator"
/gene="rrnB"
/note="rrnB T2"
regulatory complement(1875..1918)
/regulatory_class="terminator"
/gene="rrnB"
/note="rrnB T1"
rRNA complement(1912..2041)
/gene="rrnB"
/product="5S ribosomal RNA"
misc_feature complement(2122..2186)
/note="Multiple Cloning Site (MCS) from plasmid pTrc99A;
contains the following restriction sites: NcoI, EcoRI,
SacI, KpnI, SmaI, BamHI, XbaI, SalI, PstI, and HindIII"
regulatory complement(2187..2432)
/regulatory_class="promoter"
/note="trc promoter from plasmid pTrc99A; combines the -35
region from the trp promoter and the -10 region from the
lacUV5 promoter"
regulatory complement(2192..2195)
/regulatory_class="ribosome_binding_site"
/note="from lacZ"
regulatory complement(2229..2234)
/regulatory_class="minus_10_signal"
/note="from trp promoter"
regulatory complement(2252..2257)
/regulatory_class="minus_35_signal"
/note="from lacUV5 promoter"
gene complement(2463..3649)
/gene="lacI"
/allele="q"
/note="lacIq allele has been added to render the vector
autorepressed and to expand to their host range to
non-lacIq strains"
CDS complement(2489..3571)
/gene="lacI"
/allele="q"
/note="transcriptional repressor of the lac operon"
/codon_start=1
/transl_table=11
/product="LacI repressor"
/protein_id="AEG75265.1"
/translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAE
LNYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERS
GVEACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSII
FSHEDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREG
DWSAMSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDT
EDSSCYIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNT
QTASPRALADSLMQLARQVSRLESGQ"
regulatory complement(3607..3612)
/regulatory_class="minus_10_signal"
/gene="lacI"
/allele="q"
regulatory complement(3630..3635)
/regulatory_class="minus_35_signal"
/gene="lacI"
/allele="q"
misc_feature complement(3650..3680)
/note="lacIq 5' sequence"
misc_feature complement(3681..3873)
/note="intergenic region from pTrc99A"
misc_feature 3980..4920
/note="replication region from pLoxGen4 plasmid; all
elements needed for double strand DNA replication are
included"
rep_origin 4284..4292
gene complement(5049..5979)
/gene="bla"
CDS complement(5049..5909)
/gene="bla"
/function="confers resistance to ampicillin"
/codon_start=1
/transl_table=11
/product="penicillin beta-lactamase"
/protein_id="AEG75267.1"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY
IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVE
YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL
DRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL
LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA
EIGASLIKHW"
regulatory complement(5951..5956)
/regulatory_class="minus_10_signal"
/gene="bla"
regulatory complement(5974..5979)
/regulatory_class="minus_35_signal"
/gene="bla"
BASE COUNT 1530 a 1639 c 1634 g 1492 t
ORIGIN
1 aaattgtaaa cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
61 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagcccgaga
121 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
181 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccca
241 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
301 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
361 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
421 cacccgccgc gcttaatgcg ccgctacagg gcgcgtacta tggttgcttt gacgtatgcg
481 gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc attcgccatt
541 caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct
601 ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt tttcccagtc
661 acgacgttgt aaaacgacgg ccagtgccaa gctagcttaa ggtgcacggc ccacgtggcc
721 actagtactt ctcgaggtcg acggtatcga taagctggat ccataacttc gtataatgta
781 tgctatacga agttatctag cgcgtcggcc gggaagccga tctcggcttg aacgaattgt
841 taggtggcgg tacttgggtc gatatcaaag tgcatcactt cttcccgtat gcccaacttt
901 gtatagagag ccactgcggg atcgtcaccg taatctgctt gcacgtagat cacataagca
961 ccaagcgcgt tggcctcatg cttgaggaga ttgatgagcg cggtggcaat gccctgcctc
1021 cggtgctcgc cggagactgc gagatcatag atatagatct cactacgcgg ctgctcaaac
1081 ctgggcagaa cgtaagccgc gagagcgcca acaaccgctt cttggtcgaa ggcagcaagc
1141 gcgatgaatg tcttactacg gagcaagttc ccgaggtaat cggagtccgg ctgatgttgg
1201 gagtaggtgg ctacgtctcc gaactcacga ccgaaaagat caagagcagc ccgcatggat
1261 ttgacttggt cagggccgag cctacatgtg cgaatgatgc ccatacttga gccacctaac
1321 tttgttttag ggcgactgcc ctgctgcgta acatcgttgc tgctgcgtaa catcgttgct
1381 gctccataac atcaaacatc gacccacggc gtaacgcgct tgctgcttgg atgcccgagg
1441 catagactgt acaaaaaaac agtcataaca agccatgaaa accgccactg cgccgttacc
1501 accgctgcgt tcggtcaagg ttctggacca gttgcgtgag cgcatacgct acttgcatta
1561 cagtttacga accgaacagg cttatgtcga cgcgctagat aacttcgtat aatgtatgct
1621 atacgaagtt atgcggccgc gggttattgt ctcatgagcg gatacatatt tgaatgtatt
1681 tagaaaaata aacaaaaaga gtttgtagaa acgcaaaaag gccatccgtc aggatggcct
1741 tctgcttaat ttgatgcctg gcagtttatg gcgggcgtcc tgcccgccac cctccgggcc
1801 gttgcttcgc aacgttcaaa tccgctcccg gcggatttgt cctactcagg agagcgttca
1861 ccgacaaaca acagataaaa cgaaaggccc agtctttcga ctgagccttt cgttttattt
1921 gatgcctggc agttccctac tctcgcatgg ggagacccca cactaccatc ggcgctacgg
1981 cgtttcactt ctgagttcgg catggggtca ggtgggacca ccgcgctact gccgccaggc
2041 aaattctgtt ttatcagacc gcttctgcgt tctgatttaa tctgtatcag gctgaaaatc
2101 ttctctcatc cgccaaaaca gccaagcttg catgcctgca ggtcgactct agaggatccc
2161 cgggtaccga gctcgaattc catggtctgt ttcctgtgtg aaattgttat ccgctcacaa
2221 ttccacacat tatacgagcc ggatgattaa ttgtcaacag ctcatttcag aatatttgcc
2281 agaaccgtta tgatgtcggc gcaaaaaaca ttatccagaa cgggagtgcg ccttgagcga
2341 cacgaattat gcagtgattt acgacctgca cagccatacc acagcttccg atggctgcct
2401 gacgccagaa gcattggtgc accgtgcagt cgatgataag ctgtcaaacc agatcaattc
2461 gcgctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
2521 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
2581 ccagggtggt ttttcttttc accagtgaga cgggcaacag ctgattgccc ttcaccgcct
2641 ggccctgaga gagttgcagc aagcggtcca cgctggtttg ccccagcagg cgaaaatcct
2701 gtttgatggt ggttaacggc gggatataac atgagctgtc ttcggtatcg tcgtatccca
2761 ctaccgagat atccgcacca acgcgcagcc cggactcggt aatggcgcgc attgcgccca
2821 gcgccatctg atcgttggca accagcatcg cagtgggaac gatgccctca ttcagcattt
2881 gcatggtttg ttgaaaaccg gacatggcac tccagtcgcc ttcccgttcc gctatcggct
2941 gaatttgatt gcgagtgaga tatttatgcc agccagccag acgcagacgc gccgagacag
3001 aacttaatgg gcccgctaac agcgcgattt gctggtgacc caatgcgacc agatgctcca
3061 cgcccagtcg cgtaccgtct tcatgggaga aaataatact gttgatgggt gtctggtcag
3121 agacatcaag aaataacgcc ggaacattag tgcaggcagc ttccacagca atggcatcct
3181 ggtcatccag cggatagtta atgatcagcc cactgacgcg ttgcgcgaga agattgtgca
3241 ccgccgcttt acaggcttcg acgccgcttc gttctaccat cgacaccacc acgctggcac
3301 ccagttgatc ggcgcgagat ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca
3361 gactggaggt ggcaacgcca atcagcaacg actgtttgcc cgccagttgt tgtgccacgc
3421 ggttgggaat gtaattcagc tccgccatcg ccgcttccac tttttcccgc gttttcgcag
3481 aaacgtggct ggcctggttc accacgcggg aaacggtctg ataagagaca ccggcatact
3541 ctgcgacatc gtataacgtt actggtttca cattcaccac cctgaattga ctctcttccg
3601 ggcgctatca tgccataccg cgaaaggttt tgcaccattc gatggtgtca acgtaaatgc
3661 atgccgcttc gccttcgcgc gcgaattgat ctgctgcctc gcgcgtttcg gtgatgacgg
3721 tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc
3781 cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggcgcagc
3841 catgacccag tcacgtagcg atagcggagt gtaaattcgt aatcatggtc atagctgttt
3901 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
3961 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
4021 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
4081 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
4141 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
4201 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
4261 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
4321 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
4381 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
4441 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
4501 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
4561 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
4621 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
4681 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag gacagtattt
4741 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
4801 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
4861 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
4921 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
4981 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
5041 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
5101 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
5161 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
5221 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
5281 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
5341 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
5401 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
5461 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
5521 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
5581 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
5641 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
5701 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
5761 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
5821 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
5881 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
5941 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
6001 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga aaccattatt
6061 atcatgacat taacctataa aaataggcgt atcacgaggc cctttcgtct cgcgcgtttc
6121 ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg agacggtcac agcttgtctg
6181 taagcggatg ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt
6241 cggggctggc ttaactatgc ggcatcagag cagattgtac tgagagtgca ccata
//