pMAL-p5e

保存条件 ：-20℃

保质期 ：6个月

英文名 ：pMAL-p5e

库存 ：大量

供应商 ：钰博生物

规格 ：1ug/5ug

pMAL-p5e

载体基本信息

出品公司:	Ybscience
载体名称:	pMAL-p5e
质粒类型:	大肠杆菌表达载体
高拷贝/低拷贝:	--
启动子:	--
克隆方法:	多克隆位点，限制性内切酶
载体大小:	5755bp
5' 测序引物及序列:	--
3' 测序引物及序列:	--
载体标签:	--
载体抗性:	Ampicillin
筛选标记:	--
备注:	--
稳定性:	--
组成型:	--
病毒/非病毒:	--

 

载体质粒图谱和多克隆位点信息

 

载体简介

 

The vector pMAL-p5E is designed to produce maltose-binding protein (MBP) fusions, where the protein of interest can be cleaved from MBP with the specific protease Enterokinase (NEB #P8070). MBP fusions made with this vector include an N-terminal signal sequence, so the fusion protein is directed to the periplasm. The MBP has been engineered for tigher binding to amylose resin.A gene or open reading frame is inserted into a restriction site of the vector polylinker, in the same translational reading frame as the malE gene (encoding MBP). The fusion protein produced from the vector can be purified by amylose affinity chromatography. The sequence coding for the five amino acids Asp-Asp-Asp-Asp-Lys is present just upstream of the KpnI site. This allows the protein of interest to be cleaved from maltose-binding protein with enterokinase. pMAL-p5E cut with KpnI followed by treatment with the Quick Blunting Kit (NEB #E1201) produces a blunt end at the lysine codon. This allows blunt-end cloning of an insert where the first three nucleotides code for the first amino acid of the protein of interest, and enterokinase cleavage of the fusion produces a protein with no vector-derived amino acids.

 

载体序列

LOCUS       pMAL-p5E                5755 bp    DNA     circular     10-NOV-2008DEFINITION  Cloning vector pMAL-p5E, complete sequence.ACCESSIONVERSIONKEYWORDS    .SOURCE      Cloning vector pMAL-p5E  ORGANISM  Cloning vector pMAL-p5E            other sequences; artificial sequences; vectors.REFERENCE   1  (bases 1 to 5755)  AUTHORS   Riggs,P.  TITLE     Expression and purification of maltose-binding protein fusions  JOURNAL   (in) Asubel,F.A., Brent,R., Kingston,R.E., Moore,D.D.,            Seidman,J.G., Smith,J.A. and Struhl,K. (Eds.);            CURRENT PROTOCOLS IN MOLECULAR BIOLOGY: 1-14;            Greene Publishing and Wiley Interscience, New York, NY (1992)REFERENCE   2  (bases 1 to 5755)  AUTHORS   Riggs,P.D.  TITLE     Direct Submission  JOURNAL   Submitted (10-NOV-2008) Research Department, New England Biolabs,            240 County Road, Ipswich, MA 01938, USAFEATURES             Location/Qualifiers     source          1..5755                     /organism="Cloning vector pMAL-p5E"                     /mol_type="other DNA"     gene            15..1163                     /gene="lacIq"     -35_signal      15..19                     /gene="lacIq"                     /note="lacIq promoter (clockwise); TGGTG"     -10_signal      40..46                     /gene="lacIq"                     /note="lacIq promoter (clockwise); CATGATA"     CDS             81..1163                     /gene="lacIq"                     /codon_start=1                     /transl_table=11                     /product="lac repressor (LacI)"                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAE                     LNYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERS                     GVEACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSII                     FSHEDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREG                     DWSAMSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDT                     EDSSCYIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNT                     QTASPRALADSLMQLARQVSRLESGQ"     gene            1406..3046                     /gene="malE expression cassette"     -35_signal      1406..1411                     /gene="malE expression cassette"                     /note="Ptac promoter (clockwise); TTGACA"     -10_signal      1428..1433                     /gene="malE expression cassette"                     /note="Ptac promoter (clockwise); TATAAT"     CDS             1528..2832                     /gene="malE expression cassette"                     /note="malE sequence 1528..2703; spacer and enterokinase                     site 2704..2765; multiple cloning site region 2766..2832"                     /codon_start=1                     /product="maltose binding protein (MBP) fusion"                     /translation="MKIKTGARILALSALTTMMFSASALAKIEEGKLVIWINGDKGYN                     GLAEVGKKFEKDTGIKVTVEHPDKLEEKFPQVAATGDGPDIIFWAHDRFGGYAQSGLL                     AEITPDKAFQDKLYPFTWDAVRYNGKLIAYPIAVEALSLIYNKDLLPNPPKTWEEIPA                     LDKELKAKGKSALMFNLQEPYFTWPLIAADGGYAFKYENGKYDIKDVGVDNAGAKAGL                     TFLVDLIKNKHMNADTDYSIAEAAFNKGETAMTINGPWAWSNIDTSKVNYGVTVLPTF                     KGQPSKPFVGVLSAGINAASPNKELAKEFLENYLLTDEGLEAVNKDKPLGAVALKSYE                     EELVKDPRIAATMENAQKGEIMPNIPQMSAFWYAVRTAVINAASGRQTVDEALKDAQT                     NSSSNNNNNNNNNNLGDDDDKVPHMSMGGRDIVDGSEFPAGN"     primer_bind     2662..2685                     /note="malE primer sequence (NEB #S1237S)"     misc_feature    2766..2825                     /gene="malE expression cassette"                     /note="multiple cloning site (KpnI-SbfI)"     terminator      2844..2887                     /gene="malE expression cassette"                     /note="rrnB T1 transcription terminator"     primer_bind     complement(2901..2924)                     /note="MCS reverse primer sequence (NEB #SxxxxS)"     terminator      3019..3046                     /gene="malE expression cassette"                     /note="rrnB T2 transcription terminator"     gene            3095..4025                     /gene="bla"     -35_signal      3095..3100                     /gene="bla"                     /note="bla promoter (clockwise); TTCAAA"     -10_signal      3118..3123                     /gene="bla"                     /note="bla promoter (clockwise); GACAAT"     CDS             3165..4025                     /gene="bla"                     /note="ampR (confers resistance to ampicillin)"                     /codon_start=1                     /product="beta-lactamase"                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY                     IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVE                     YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL                     DRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL                     LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA                     EIGASLIKHW"     rep_origin      4113..4701                     /note="pMB1 origin of replication (clockwise) (RNAII -35                     to RNA/DNA switch point)"     gene            complement(5071..5262)                     /gene="rop"     CDS             complement(5071..5262)                     /gene="rop"                     /codon_start=1                     /transl_table=11                     /product="ROP protein"                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDH                     ADELYRSCLARFGDDGENL"BASE COUNT     1439 a   1455 c   1538 g   1323 tORIGIN        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 tatttcggta 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 taagttagct 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 gcaattgacc aacaaggacc atagattatg aaaataaaaa caggtgcacg catcctcgca     1561 ttatccgcat taacgacgat gatgttttcc gcctcggctc tcgccaaaat cgaagaaggt     1621 aaactggtaa tctggattaa cggcgataaa ggctataacg gtctcgctga agtcggtaag     1681 aaattcgaga aagataccgg aattaaagtc accgttgagc atccggataa actggaagag     1741 aaattcccac aggttgcggc aactggcgat ggccctgaca ttatcttctg ggcacacgac     1801 cgctttggtg gctacgctca atctggcctg ttggctgaaa tcaccccgga caaagcgttc     1861 caggacaagc tgtatccgtt tacctgggat gccgtacgtt acaacggcaa gctgattgct     1921 tacccgatcg ctgttgaagc gttatcgctg atttataaca aagatctgct gccgaacccg     1981 ccaaaaacct gggaagagat cccggcgctg gataaagaac tgaaagcgaa aggtaagagc     2041 gcgctgatgt tcaacctgca agaaccgtac ttcacctggc cgctgattgc tgctgacggg     2101 ggttatgcgt tcaagtatga aaacggcaag tacgacatta aagacgtggg cgtggataac     2161 gctggcgcga aagcgggtct gaccttcctg gttgacctga ttaaaaacaa acacatgaat     2221 gcagacaccg attactccat cgcagaagct gcctttaata aaggcgaaac agcgatgacc     2281 atcaacggcc cgtgggcatg gtccaacatc gacaccagca aagtgaatta tggtgtaacg     2341 gtactgccga ccttcaaggg tcaaccatcc aaaccgttcg ttggcgtgct gagcgcaggt     2401 attaacgccg ccagtccgaa caaagagctg gcaaaagagt tcctcgaaaa ctatctgctg     2461 actgatgaag gtctggaagc ggttaataaa gacaaaccgc tgggtgccgt agcgctgaag     2521 tcttacgagg aagagttggt gaaagatccg cgtattgccg ccactatgga aaacgcccag     2581 aaaggtgaaa tcatgccgaa catcccgcag atgtccgctt tctggtatgc cgtgcgtact     2641 gcggtgatca acgccgccag cggtcgtcag actgtcgatg aagccctgaa agacgcgcag     2701 actaattcga gctcgaacaa caacaacaat aacaataaca acaacctcgg ggatgacgat     2761 gacaaggtac cgcatatgtc catgggcggc cgcgatatcg tcgacggatc cgaattccct     2821 gcaggtaatt aaataagctt caaataaaac gaaaggctca gtcgaaagac tgggcctttc     2881 gttttatctg ttgtttgtcg gtgaacgctc tcctgagtag gacaaatccg ccgggagcgg     2941 atttgaacgt tgcgaagcaa cggcccggag ggtggcgggc aggacgcccg ccataaactg     3001 ccaggcatca aattaagcag aaggccatcc tgacggatgg cctttttgcg tttctacaaa     3061 ctctttcggt ccgttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac     3121 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt     3181 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag     3241 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg     3301 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgtttcccaa     3361 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtgtt gacgccgggc     3421 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag     3481 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa     3541 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc     3601 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg     3661 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa     3721 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa     3781 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg     3841 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag     3901 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg     3961 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt     4021 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttcctt aggactgagc     4081 gtcaaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc     4141 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag     4201 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc     4261 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac     4321 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc     4381 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt     4441 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt     4501 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc     4561 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt     4621 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca     4681 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt     4741 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt     4801 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag     4861 tcagtgagcg aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc     4921 ggtatttcac accgcatata aggtgcactg tgactgggtc atggctgcgc cccgacaccc     4981 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca     5041 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg     5101 cgcgaggcag ctgcggtaaa gctcatcagc gtggtcgtgc agcgattcac agatgtctgc     5161 ctgttcatcc gcgtccagct cgttgagttt ctccagaagc gttaatgtct ggcttctgat     5221 aaagcgggcc atgttaaggg cggttttttc ctgtttggtc actgatgcct ccgtgtaagg     5281 gggatttctg ttcatggggg taatgatacc gatgaaacga gagaggatgc tcacgatacg     5341 ggttactgat gatgaacatg cccggttact ggaacgttgt gagggtaaac aactggcggt     5401 atggatgcgg cgggaccaga gaaaaatcac tcagggtcaa tgccagcgct tcgttaatac     5461 agatgtaggt gttccacagg gtagccagca gcatcctgcg atgcagatcc ggaacataat     5521 ggtgcagggc gctgacttcc gcgtttccag actttacgaa acacggaaac cgaagaccat     5581 tcatgttgtt gctcaggtcg cagacgtttt gcagcagcag tcgcttcacg ttcgctcgcg     5641 tatcggtgat tcattctgct aaccagtaag gcaaccccgc cagcctagcc gggtcctcaa     5701 cgacaggagc acgatcatgc gcacccgtgg ccaggaccca acgctgcccg aaatt