LOCUS pCYB3 6846 bp DNA circular 10-DEC-2007
DEFINITION Cloning vector pCYB3, complete sequence.
ACCESSION
VERSION
KEYWORDS .
SOURCE Cloning vector pCYB3
ORGANISM Cloning vector pCYB3
other sequences; artificial sequences; vectors.
REFERENCE 1 (bases 1 to 6846)
AUTHORS Chong,S., Mersha,F.B., Comb,D.G., Scott,M.E., Landry,D.,
Vence,L.M., Perler,F.B., Benner,J., Kucera,R.B., Hirvonen,C.A.,
Pelletier,J.J., Paulus,H. and Xu,M.-Q.
TITLE Single-column purification of free recombinant proteins using a
self-cleavable affinity tag derived from a protein splicing element
JOURNAL Gene 192, 271-281 (1997)
PUBMED 9224900
REFERENCE 2 (bases 1 to 6846)
AUTHORS Perler,F.B.
TITLE InBase, the intein database
JOURNAL Nucleic Acids Res 30 (1), 383-384 (2002)
PUBMED 11752343
REFERENCE 3 (bases 1 to 6846)
AUTHORS Ghosh,I. and Xu,M.Q.
TITLE Direct Submission
JOURNAL Submitted (10-DEC-2007) Research Department, New England Biolabs,
240 County Road, Ipswich, MA 01938, USA
FEATURES Location/Qualifiers
source 1..6846
/organism="Cloning vector pCYB3"
/mol_type="other DNA"
gene 15..1163
/gene="lacIq"
promoter 15..46
/gene="lacIq"
/note="lacIq promoter (overexpressing mutant) (clockwise)"
CDS 81..1163
/gene="lacIq"
/exception="alternative start codon"
/codon_start=1
/transl_table=11
/product="lac repressor (LacI)"
/translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAE
LNYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERS
GVEACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSII
FSHEDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREG
DWSAMSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDT
EDSSCYIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNT
QTASPRALADSLMQLARQVSRLESGQ"
promoter 1406..1433
/note="Ptac promoter (clockwise)"
protein_bind 1438..1462
/note="lac operator"
/bound_moiety="LacI"
primer_bind 1446..1469
/note="Ptac forward sequencing primer (NEB #1260,
discontinued)"
RBS 1515..1518
/note="RBS for expression CDS"
misc_feature 1526..1575
/gene="expression CDS"
/note="multiple cloning site (NcoI-SapI)"
gene 1528..3129
/gene="expression CDS"
CDS 1528..3129
/gene="expression CDS"
/note="multiple cloning site (NcoI-SapI) 1526..1575; Sce
VMA intein sequence 1576..2937; chitin binding domain
(CBD) sequence 2971..3129"
/codon_start=1
/product="intein/CBD fusion"
/translation="MASSRVDSREFLEGSSCFAKGTNVLMADGSIECIENIEVGNKVM
GKDGRPREVIKLPRGRETMYSVVQKSQHRAHKSDSSREVPELLKFTCNATHELVVRTP
RSVRRLSRTIKGVEYFEVITFEMGQKKAPDGRIVELVKEVSKSYPISEGPERANELVE
SYRKASNKAYFEWTIEARDLSLLGSHVRKATYQTYAPILYENDHFFDYMQKSKFHLTI
EGPKVLAYLLGLWIGDGLSDRATFSVDSRDTSLMERVTEYAEKLNLCAEYKDRKEPQV
AKTVNLYSKVVRGNGIRNNLNTENPLWDAIVGLGFLKDGVKNIPSFLSTDNIGTRETF
LAGLIDSDGYVTDEHGIKATIKTIHTSVRDGLVSLARSLGLVVSVNAEPAKVDMNVTK
HKISYAIYMSGGDVLLNVLSKCAGSKKFRPAPAAAFARECRGFYFELQELKEDDYYGI
TLSDDSDHQFLLGSQVVVHACGGLTGLNSGLTTNPGVSAWQVNTAYTAGQLVTYNGKT
YKCLQPHTSLAGWEPSNVPALWQLQ"
primer_bind complement(1639..1662)
/note="intein reverse primer sequence (NEB #1261S)"
gene 3677..4537
/gene="bla"
CDS 3677..4537
/gene="bla"
/note="ampR (confers resistance to ampicillin)"
/codon_start=1
/product="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY
IELDLNSGKILESFRPEERSPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVE
YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL
DRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL
LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA
EIGASLIKHW"
sig_peptide 3677..3745
/gene="bla"
/note="required for secretion to the periplasm; cleaved
off to form the mature beta-lactamase protein"
rep_origin 4579..5092
/note="M13 origin of replication (+ -)"
rep_origin 5203..5791
/note="pMB1 origin of replication (clockwise) (RNAII -35
to RNA/DNA switch point)"
gene complement(6160..6351)
/gene="rop"
CDS complement(6160..6351)
/gene="rop"
/exception="alternative start codon"
/codon_start=1
/transl_table=11
/product="ROP protein"
/translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDH
ADELYRSCLARFGDDGENL"
BASE COUNT 1736 a 1623 c 1779 g 1708 t
ORIGIN
1 ccgacaccat cgaatggtgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga
61 gtcaattcag ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg
121 gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa
181 cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac
241 aacaactggc gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc
301 acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg
361 tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc
421 ttctcgcgca acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca
481 ttgctgtgga agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga
541 cacccatcaa cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc
601 tggtcgcatt gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg
661 cgcgtctgcg tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag
721 cggaacggga aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga
781 atgagggcat cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa
841 tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga tatctcggta gtgggatacg
901 acgataccga agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc
961 gcctgctggg gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga
1021 agggcaatca gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata
1081 cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt
1141 cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct cactcattag
1201 gcacaattct catgtttgac agcttatcat cgactgcacg gtgcaccaat gcttctggcg
1261 tcaggcagcc atcggaagct gtggtatggc tgtgcaggtc gtaaatcact gcataattcg
1321 tgtcgctcaa ggcgcactcc cgttctggat aatgtttttt gcgccgacat cataacggtt
1381 ctggcaaata ttctgaaatg agctgttgac aattaatcat cggctcgtat aatgtgtgga
1441 attgtgagcg gataacaatt tcacacagga aacagccagt ccgtttaggt gttttcacga
1501 gcacttcacc aacaaggacc atagaccatg gctagctcgc gagtcgactc tagagaattc
1561 ctcgagggct cttcctgctt tgccaagggt accaatgttt taatggcgga tgggtctatt
1621 gaatgtattg aaaacattga ggttggtaat aaggtcatgg gtaaagatgg cagacctcgt
1681 gaggtaatta aattgcccag aggaagagaa actatgtaca gcgtcgtgca gaaaagtcag
1741 cacagagccc acaaaagtga ctcaagtcgt gaagtgccag aattactcaa gtttacgtgt
1801 aatgcgaccc atgagttggt tgttagaaca cctcgtagtg tccgccgttt gtctcgtacc
1861 attaagggtg tcgaatattt tgaagttatt acttttgaga tgggccaaaa gaaagccccc
1921 gacggtagaa ttgttgagct tgtcaaggaa gtttcaaaga gctacccaat atctgagggg
1981 cctgagagag ccaacgaatt agtagaatcc tatagaaagg cttcaaataa agcttatttt
2041 gagtggacta ttgaggccag agatctttct ctgttgggtt cccatgttcg taaagctacc
2101 taccagactt acgctccaat tctttatgag aatgaccact ttttcgacta catgcaaaaa
2161 agtaagtttc atctcaccat tgaaggtcca aaagtacttg cttatttact tggtttatgg
2221 attggtgatg gattgtctga cagggcaact ttttcggttg attccagaga tacttctttg
2281 atggaacgtg ttactgaata tgctgaaaag ttgaatttgt gcgccgagta taaggacaga
2341 aaagaaccac aagttgccaa aactgttaat ttgtactcta aagttgtcag aggtaatggt
2401 attcgcaata atcttaatac tgagaatcca ttatgggacg ctattgttgg cttaggattc
2461 ttgaaggacg gtgtcaaaaa tattccttct ttcttgtcta cggacaatat cggtactcgt
2521 gaaacatttc ttgctggtct aattgattct gatggctatg ttactgatga gcatggtatt
2581 aaagcaacaa taaagacaat tcatacttct gtcagagatg gtttggtttc ccttgctcgt
2641 tctttaggct tagtagtctc ggttaacgca gaacctgcta aggttgacat gaatgtcacc
2701 aaacataaaa ttagttatgc tatttatatg tctggtggag atgttttgct taacgttctt
2761 tcgaagtgtg ccggctctaa aaaattcagg cctgctcccg ccgctgcttt tgcacgtgag
2821 tgccgcggat tttatttcga gttacaagaa ttgaaggaag acgattatta tgggattact
2881 ttatctgatg attctgatca tcagtttttg cttggatccc aggttgtcgt ccatgcatgc
2941 ggtggcctga ccggtctgaa ctcaggcctc acgacaaatc ctggtgtatc cgcttggcag
3001 gtcaacacag cttatactgc gggacaattg gtcacatata acggcaagac gtataaatgt
3061 ttgcagcccc acacctcctt ggcaggatgg gaaccatcca acgttcctgc cttgtggcag
3121 cttcaatgac tgcagcctga atggcgaatg gcagcttggc tgttttggcg gatgagataa
3181 gattttcagc ctgatacaga ttaaatcaga acgcagaagc ggtctgataa aacagaattt
3241 gcctggcggc agtagcgcgg tggtcccacc tgaccccatg ccgaactcag aagtgaaacg
3301 ccgtagcgcc gatggtagtg tggggtctcc ccatgcgaga gtagggaact gccaggcatc
3361 aaataaaacg aaaggctcag tcgaaagact gggcctttcg ttttatctgt tgtttgtcgg
3421 tgaacgctct cctgagtagg acaaatccgc cgggagcgga tttgaacgtt gcgaagcaac
3481 ggcccggagg gtggcgggca ggacgcccgc cataaactgc caggcatcaa attaagcaga
3541 aggccatcct gacggatggc ctttttgcgt ttctacaaac tctttttgtt tatttttcta
3601 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
3661 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
3721 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
3781 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
3841 tgagagtttt cgccccgaag aacgttctcc aatgatgagc acttttaaag ttctgctatg
3901 tggcgcggta ttatcccgtg ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
3961 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
4021 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
4081 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
4141 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
4201 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgtaaactat taactggcga
4261 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
4321 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
4381 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
4441 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
4501 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
4561 tatactttag attgatttac cccggttgat aatcagaaaa gccccaaaaa caggaagatt
4621 gtataagcaa atatttaaat tgtaaacgtt aatattttgt taaaattcgc gttaaatttt
4681 tgttaaatca gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca
4741 aaagaatagc ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta
4801 aagaacgtgg actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta
4861 cgtgaaccat cacccaaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg
4921 aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga
4981 aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg
5041 ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtaaaaggat
5101 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
5161 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
5221 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
5281 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
5341 aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
5401 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
5461 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
5521 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
5581 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
5641 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
5701 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
5761 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
5821 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt
5881 ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga
5941 gcgcagcgag tcagtgagcg aggaagctat ggtgcactct cagtacaatc tgctctgatg
6001 ccgcatagtt aagccagtat acactccgct atcgctacgt gactgggtca tggctgcgcc
6061 ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc
6121 ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc
6181 accgaaacgc gcgaggcagc tgcggtaaag ctcatcagcg tggtcgtgca gcgattcaca
6241 gatgtctgcc tgttcatccg cgtccagctc gttgagtttc tccagaagcg ttaatgtctg
6301 gcttctgata aagcgggcca tgttaagggc ggttttttcc tgtttggtca cttgatgcct
6361 ccgtgtaagg gggaatttct gttcatgggg gtaatgatac cgatgaaacg agagaggatg
6421 ctcacgatac gggttactga tgatgaacat gcccggttac tggaacgttg tgagggtaaa
6481 caactggcgg tatggatgcg gcgggaccag agaaaaatca ctcagggtca atgccagcgc
6541 ttcgttaata cagatgtagg tgttccacag ggtagccagc agcatcctgc gatgcagatc
6601 cggaacataa tggtgcaggg cgctgacttc cgcgtttcca gactttacga aacacggaaa
6661 ccgaagacca ttcatgttgt tgctcaggtc gcagacgttt tgcagcagca gtcgcttcac
6721 gttcgctcgc gtatcggtga ttcattctgc taaccagtaa ggcaaccccg ccagcctagc
6781 cgggtcctca acgacaggag cacgatcatg cgcacccgtg gccaggaccc aacgctgccc
6841 gaaatt
//