| 质粒名称: | pHEA |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 168297 |
| 存储实验室: | Luis Ángel Fernández |
|
载体骨架基本信息 |
|
| 载体名称: | pAK-Not |
| 载体出品公司: | Luis Angel Fernández |
| 空载体大小: | 3735 bp |
| 完整质粒大小: | 4974 bp |
| 载体修饰: | -- |
| 载体类型: | 细菌表达 |
| 筛选标记: | -- |
| 载体抗性: | 氯霉素25 μg/mL |
| 生长温度: | 37 ℃ |
| 宿主菌: | DH5a |
| 拷贝数: | 高拷贝 |
|
插入基因信息 |
|
| 插入基因名称: | Enterohemorraghic E. coli autotransporter EhaA C-terminal fragment (C-EhaA) |
| 物种来源: | Escherichia coli |
| 插入大小: | 1239 bp |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子: | LacI-Plac |
| 标签/融合蛋白: |
PelB signal peptide（插入物上的N端）
6xHis（插入件上的N端子）
E-tag（插入件上的N端子）
|
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | -- |
| 3’克隆位点: | -- |
| 5’测序引物 | -- |
| 3’测序引物 | -- |
|
文献引用方法 |
|
| 材料和方法部分: | pHEA was a gift from Luis Ángel Fernández (Addgene plasmid # 168297 ; http://n2t.net/addgene:168297 ; RRID:Addgene_168297) |
| 参考文献部分: | Comparative analysis of the biochemical and functional properties of C-terminal domains of autotransporters. Marin E, Bodelon G, Fernandez LA. J Bacteriol. 2010 Nov;192(21):5588-602. doi: 10.1128/JB.00432-10. Epub 2010 Aug 27. 10.1128/JB.00432-10 PubMed 20802036 |
买家导航
载体基本信息
载体图谱质粒图谱和多克隆位点信息
载体简介
载体序列
LOCUS pHEA 4974 bp ds-DNA circular SYN 09-JUN-2021
DEFINITION Bacterial surface display of polypeptides (nanobodies) in E. coli.
ACCESSION .
VERSION .
KEYWORDS pHEA
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4974)
AUTHORS Marin E, Bodelon G, Fernandez LA
TITLE Comparative analysis of the biochemical and functional properties of
C-terminal domains of autotransporters.
JOURNAL J Bacteriol. 2010 Nov;192(21):5588-602. doi: 10.1128/JB.00432-10.
Epub 2010 Aug 27.
PUBMED 20802036
REFERENCE 2 (bases 1 to 4974)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Jun 9, 2021 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..4974
/organism="synthetic DNA construct"
/mol_type="other DNA"
protein_bind 591..612
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 627..657
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 665..681
/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 670..692
/label=M13/pUC Reverse
/note="In lacZ gene"
primer_bind 689..705
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 689..705
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
sig_peptide 741..806
/gene="pelB (fragment)"
/product="leader peptide for secretion"
/label=pelB signal sequence
CDS 807..824
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
rep_origin 2051..2506
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(2138..2157)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 2348..2369
/label=F1ori-F
/note="F1 origin, forward primer"
CDS 2693..3352
/codon_start=1
/gene="cat"
/product="chloramphenicol acetyltransferase"
/label=CmR
/note="confers resistance to chloramphenicol"
/translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
primer_bind complement(2759..2778)
/label=CAT-R
/note="Chloramphenicol resistance gene, reverse primer"
rep_origin 3684..4272
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 4173..4192
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
promoter 4337..4414
/gene="lacI"
/label=lacI promoter
CDS 4415..523
/codon_start=1
/gene="lacI"
/product="lac repressor"
/label=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)."
/translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
ALADSLMQLARQVSRLESGQ"
primer_bind complement(4434..4453)
/label=LacI-R
/note="LacI, reverse primer"
ORIGIN
1 cccattaagt tctgtctcgg cgcgtctgcg tctggctggc tggcataaat atctcactcg
61 caatcaaatt cagccgatag cggaacggga aggcgactgg agtgccatgt ccggttttca
121 acaaaccatg caaatgctga atgagggcat cgttcccact gcgatgctgg ttgccaacga
181 tcagatggcg ctgggcgcaa tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga
241 catctcggta gtgggatacg acgataccga agacagctca tgttatatcc cgccgttaac
301 caccatcaaa caggattttc gcctgctggg gcaaaccagc gtggaccgct tgctgcaact
361 ctctcagggc caggcggtga agggcaatca gctgttgccc gtctcactgg tgaaaagaaa
421 aaccaccctg gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat
481 gcagctggca cgacaggttt cccgactgga aagcgggcag tgagcggtac ccgataaaag
541 cggcttcctg acaggaggcc gttttgtttt gcagcccacc tcaacgcaat taatgtgagt
601 tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt
661 ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga ttacgaattt
721 ctagataacg agggcaaatc atgaaatacc tattgcctac ggcagccgct ggattgttat
781 tactcgcggc ccagccggcc atggcgcatc accatcacca tcacgcggcc gctgcggggg
841 ccgaattcgt cgacggtgcg ccggtgccgt atccggatcc gctggaaccg atcgacaatt
901 cagccgcaat tagtatggca aatccacgtc caccaacacc gcggacacct acgccgggtc
961 cggatctgaa tgtggataat gacctgcgac cggaggcggg tagctacatt gcgaaccttg
1021 cagcagcgaa taccatgttc accacgcgtc tgcatgagcg tctgggtaat acgtactata
1081 ccgacatggt gacgggtgag cagaaacaaa ccactatgtg gatgcgccat gaaggtggtc
1141 ataataaatg gcgtgatggc agcggccagc tgaaaaccca aagcaatcgc tatgttctgc
1201 aactgggagg cgatgtcgcg cagtggagcc aaaacggcag cgaccgctgg catgttgggg
1261 tcatggcggg atatggcaac agcgacagca aaaccatttc ctcgcgaacc ggttatcgtg
1321 caaaagcgag tgtgaacgga tatagcacag gcctctatgc cacctggtat gccgatgacg
1381 agtcgcgtaa tggcgcgtat ctcgacagtt gggcgcagta cagctggttt gataacacag
1441 tgaaagggga tgacttacaa agtgaatcct ataaatcaaa aggatttacc gcttcactgg
1501 aagctggata caaacacaaa ttagctgaat ttaatggcag ccagggaacg cgtaatgaat
1561 ggtatgttca gccgcaagca caggttacct ggatgggagt caaagccgat aagcaccgcg
1621 aaagcaacgg aaccctcgtt catagcaacg gtgatggcaa tgttcaaacc cgacttggcg
1681 taaaaacctg gctgaagagc caccataaaa tggatgacgg taaatcccgc gagttccagc
1741 cgtttgtaga agtgaactgg ctacataaca gtaaggattt cagcaccagt atggatggcg
1801 tgtctgtcac tcaggatgga gcccgaaata ttgctgagat aaaaaccggg gtggaaggac
1861 agctaaatgc caacctgaat gtctggggga atgtgggcgt tcaggttgcc gataggggat
1921 ataatgacac ctctgcaatg gttggcatta agtggcaatt ctgaaagctt gacctgtgaa
1981 gtgaaaaatg gcgcacattg tgcgacattt tttttgtctg ccgtttaccg ctactgcgtc
2041 acggatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
2101 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
2161 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
2221 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
2281 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
2341 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
2401 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
2461 caaaaattta acgcgaattt taacaaaata ttaacgctta caatttcagg tggcactttt
2521 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
2581 ccgctcatgt cgagacgttg ggtgaggttc caactttcac cataatgaaa taagatcact
2641 accgggcgta ttttttgagt tatcgagatt ttcaggagct aaggaagcta aaatggagaa
2701 aaaaatcact ggatatacca ccgttgatat atcccaatgg catcgtaaag aacattttga
2761 ggcatttcag tcagttgctc aatgtaccta taaccagacc gttcagctgg atattacggc
2821 ctttttaaag accgtaaaga aaaataagca caagttttat ccggccttta ttcacattct
2881 tgcccgcctg atgaatgctc atccggagtt ccgtatggca atgaaagacg gtgagctggt
2941 gatatgggat agtgttcacc cttgttacac cgttttccat gagcaaactg aaacgttttc
3001 atcgctctgg agtgaatacc acgacgattt ccggcagttt ctacacatat attcgcaaga
3061 tgtggcgtgt tacggtgaaa acctggccta tttccctaaa gggtttattg agaatatgtt
3121 tttcgtctca gccaatccct gggtgagttt caccagtttt gatttaaacg tggccaatat
3181 ggacaacttc ttcgcccccg ttttcaccat gggcaaatat tatacgcaag gcgacaaggt
3241 gctgatgccg ctggcgattc aggttcatca tgccgtttgt gatggcttcc atgtcggcag
3301 aatgcttaat gaattacaac agtactgcga tgagtggcag ggcggggcgt aattttttta
3361 aggcagttat tggtgccctt aaacgcctgg ttgctacgcc tgaataagtg ataataagcg
3421 gatgaatggc agaaattcga aagcaaattc gacccggtcg tcggttcagg gcagggtcgt
3481 taaatagccg cttatgtcta ttgctggttt accggtttat tgactaccgg aagcagtgtg
3541 accgtgtgct tctcaaatgc ctgaggccag tttgctcagg ctctccccgt ggaggtaata
3601 attgctcgaa tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
3661 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
3721 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
3781 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
3841 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
3901 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
3961 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
4021 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
4081 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
4141 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
4201 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
4261 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
4321 tttgctcaca tgacccgaca ccatcgaatg gcgcaaaacc tttcgcggta tggcatgata
4381 gcgcccggaa gagagtcaat tcagggtggt gaatgtgaaa ccagtaacgt tatacgatgt
4441 cgcagagtat gccggtgtct cttatcagac cgtttcccgc gtggtgaacc aggccagcca
4501 cgtttctgcg aaaacgcggg aaaaagtgga agcggcgatg gcggagctga attacattcc
4561 caaccgcgtg gcacaacaac tggcgggcaa acagtcgttg ctgattggcg ttgccacctc
4621 cagtctggcc ctgcacgcgc cgtcgcaaat tgtcgcggcg attaaatctc gcgccgatca
4681 actgggtgcc agcgtggtgg tgtcgatggt agaacgaagc ggcgtcgaag cctgtaaagc
4741 ggcggtgcac aatcttctcg cgcaacgcgt cagtgggctg atcattaact atccgctgga
4801 tgaccaggat gccattgctg tggaagctgc ctgcactaat gttccggcgt tatttcttga
4861 tgtctctgac cagacaccca tcaacagtat tattttctcc catgaagacg gtacgcgact
4921 gggcgtggag catctggtcg cattgggtca ccagcaaatc gcgctgttag cggg
//
