| 质粒名称: | Sender plasmid |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 127677 |
| 存储实验室: | Friedrich Simmel |
|
载体骨架基本信息 |
|
| 载体名称: | pETDuet-1 |
| 载体出品公司: | Novagen |
| 空载体大小: | 5420 bp |
| 完整质粒大小: | 5986 bp |
| 载体修饰: | -- |
| 载体类型: | 细菌表达，合成生物学 |
| 筛选标记: | -- |
| 载体抗性: | 氨苄青霉素，100μg/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: | LuxI |
| 物种来源: | V. fischeri |
| 插入大小: | 643 bp |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子: | pT7 |
| 标签/融合蛋白: | LVA（插入件上的C端子） |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | XbaI(未破坏) |
| 3’克隆位点: | HindIII (未破坏) |
| 5’测序引物 | GTGATGTCGGCGATATAGG |
| 3’测序引物 | GTTAATTAAGCTGCGCTAGTAG |
|
文献引用方法 |
|
| 材料和方法部分: | Sender plasmid was a gift from Friedrich Simmel (Addgene plasmid # 127677 ; http://n2t.net/addgene:127677 ; RRID:Addgene_127677) |
| 参考文献部分: | Chemical communication between bacteria and cell-free gene expression systems within linear chains of emulsion droplets. Schwarz-Schilling M, Aufinger L, Muckl A, Simmel FC. Integr Biol (Camb). 2016 Apr 18;8(4):564-70. doi: 10.1039/c5ib00301f. Epub 2016 Jan 18. 10.1039/c5ib00301f PubMed 26778746 |
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 Sender plasmid 5999 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Encodes the synthase LuxI for the quorum-sensing molecule C6-HSL..
ACCESSION .
VERSION .
KEYWORDS Sender plasmid
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5999)
AUTHORS Schwarz-Schilling M, Aufinger L, Muckl A, Simmel FC
TITLE Chemical communication between bacteria and cell-free gene
expression systems within linear chains of emulsion droplets.
JOURNAL Integr Biol (Camb). 2016 Apr 18;8(4):564-70. doi:
10.1039/c5ib00301f. Epub 2016 Jan 18.
PUBMED 26778746
REFERENCE 2 (bases 1 to 5999)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..5999
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 1..19
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 20..44
/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)."
CDS 651..683
/codon_start=1
/product="C-terminal peptide that mediates degradation in
bacteria through the ClpXP and ClpAP proteases (McGinness
et al., 2006)"
/label=ssrA tag (LVA)
/note="mutant LVA variant that confers accelerated
degradation under some conditions (Andersen et al., 1998)"
/translation="AANDENYALVA"
misc_feature 715..735
/label=BioBrick suffix
/note="universal suffix for all parts"
promoter 813..831
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
protein_bind 832..856
/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)."
CDS 965..1009
/codon_start=1
/product="affinity and epitope tag derived from pancreatic
ribonuclease A"
/label=S-Tag
/translation="KETAAAKFERQHMDS"
primer_bind complement(1047..1065)
/label=T7 Term
/note="T7 terminator, reverse primer"
terminator 1061..1108
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
rep_origin 1145..1600
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(1232..1251)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 1442..1463
/label=F1ori-F
/note="F1 origin, forward primer"
CDS complement(1715..2575)
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
primer_bind 2338..2357
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(2576..2668)
/gene="bla"
/label=AmpR promoter
rep_origin 2749..3337
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 3238..3257
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 3491..3508
/label=L4440
/note="L4440 vector, forward primer"
misc_feature 3523..3662
/label=bom
/note="basis of mobility region from pBR322"
primer_bind complement(3588..3607)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 3748..3770
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
CDS complement(3764..3955)
/codon_start=1
/gene="rop"
/product="Rop protein, which maintains plasmids at low copy
number"
/label=rop
/translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
DELYRSCLARFGDDGENL"
CDS complement(4527..5609)
/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 5571..5590
/label=LacI-R
/note="LacI, reverse primer"
promoter complement(5610..5687)
/gene="lacI"
/label=lacI promoter
primer_bind 5893..5912
/label=pBRrevBam
/note="pBR322 vectors, tet region, downstream of BamHI,
reverse primer"
ORIGIN
1 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gagaaagagg
61 ggaaatacta gatgactata atgataaaaa aatcggattt tttggcaatt ccatcggagg
121 agtataaagg tattctaagt cttcgttatc aagtgtttaa gcaaagactt gagtgggact
181 tagttgtaga aaataacctt gaatcagatg agtatgataa ctcaaatgca gaatatattt
241 atgcttgtga tgatactgaa aatgtaagtg gatgctggcg tttattacct acaacaggtg
301 attatatgct gaaaagtgtt tttcctgaat tgcttggtca acagagtgct cccaaagatc
361 ctaatatagt cgaattaagt cgttttgctg taggtaaaaa tagctcaaag ataaataact
421 ctgctagtga aattacaatg aaactatttg aagctatata taaacacgct gttagtcaag
481 gtattacaga atatgtaaca gtaacatcaa cagcaataga gcgattttta aagcgtatta
541 aagttccttg tcatcgtatt ggagacaaag aaattcatgt attaggtgat actaaatcgg
601 ttgtattgtc tatgcctatt aatgaacagt ttaaaaaagc agtcttaaat gctgcaaacg
661 acgaaaacta cgctttagta gcttaataac tctgatagtg ctagtgtaga tctctactag
721 tagcggccgc tgcaggtcga caagcttgcg gccgcataat gcttaagtcg aacagaaagt
781 aatcgtattg tacacggccg cataatcgaa attaatacga ctcactatag gggaattgtg
841 agcggataac aattccccat cttagtatat tagttaagta taagaaggag atatacatat
901 ggcagatctc aattggatat cggccggcca cgcgatcgct gacgtcggta ccctcgagtc
961 tggtaaagaa accgctgctg cgaaatttga acgccagcac atggactcgt ctactagcgc
1021 agcttaatta acctaggctg ctgccaccgc tgagcaataa ctagcataac cccttggggc
1081 ctctaaacgg gtcttgaggg gttttttgct gaaaggagga actatatccg gattggcgaa
1141 tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
1201 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
1261 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
1321 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt
1381 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
1441 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
1501 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
1561 tttaacgcga attttaacaa aatattaacg tttacaattt ctggcggcac gatggcatga
1621 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
1681 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
1741 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
1801 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
1861 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
1921 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
1981 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
2041 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
2101 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
2161 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
2221 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
2281 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
2341 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
2401 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
2461 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
2521 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
2581 tcctttttca atcatgattg aagcatttat cagggttatt gtctcatgag cggatacata
2641 tttgaatgta tttagaaaaa taaacaaata ggtcatgacc aaaatccctt aacgtgagtt
2701 ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt
2761 ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg
2821 tttgccggat caagagctac caactctttt tccgaaggta actggcttca gcagagcgca
2881 gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt
2941 agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga
3001 taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
3061 gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact
3121 gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga
3181 caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg
3241 aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt
3301 tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt
3361 acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga
3421 ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac
3481 gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcctgatgc ggtattttct
3541 ccttacgcat ctgtgcggta tttcacaccg caatggtgca ctctcagtac aatctgctct
3601 gatgccgcat agttaagcca gtatacactc cgctatcgct acgtgactgg gtcatggctg
3661 cgccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
3721 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
3781 catcaccgaa acgcgcgagg cagctgcggt aaagctcatc agcgtggtcg tgaagcgatt
3841 cacagatgtc tgcctgttca tccgcgtcca gctcgttgag tttctccaga agcgttaatg
3901 tctggcttct gataaagcgg gccatgttaa gggcggtttt ttcctgtttg gtcactgatg
3961 cctccgtgta agggggattt ctgttcatgg gggtaatgat accgatgaaa cgagagagga
4021 tgctcacgat acgggttact gatgatgaac atgcccggtt actggaacgt tgtgagggta
4081 aacaactggc ggtatggatg cggcgggacc agagaaaaat cactcagggt caatgccagc
4141 gcttcgttaa tacagatgta ggtgttccac agggtagcca gcagcatcct gcgatgcaga
4201 tccggaacat aatggtgcag ggcgctgact tccgcgtttc cagactttac gaaacacgga
4261 aaccgaagac cattcatgtt gttgctcagg tcgcagacgt tttgcagcag cagtcgcttc
4321 acgttcgctc gcgtatcggt gattcattct gctaaccagt aaggcaaccc cgccagccta
4381 gccgggtcct caacgacagg agcacgatca tgctagtcat gccccgcgcc caccggaagg
4441 agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc taatgagtga
4501 gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
4561 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgcc
4621 agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt caccgcctgg
4681 ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt
4741 ttgatggtgg ttaacggcgg gatataacat gagctgtctt cggtatcgtc gtatcccact
4801 accgagatgt ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc
4861 gccatctgat cgttggcaac cagcatcgca gtgggaacga tgccctcatt cagcatttgc
4921 atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc tatcggctga
4981 atttgattgc gagtgagata tttatgccag ccagccagac gcagacgcgc cgagacagaa
5041 cttaatgggc ccgctaacag cgcgatttgc tggtgaccca atgcgaccag atgctccacg
5101 cccagtcgcg taccgtcttc atgggagaaa ataatactgt tgatgggtgt ctggtcagag
5161 acatcaagaa ataacgccgg aacattagtg caggcagctt ccacagcaat ggcatcctgg
5221 tcatccagcg gatagttaat gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc
5281 gccgctttac aggcttcgac gccgcttcgt tctaccatcg acaccaccac gctggcaccc
5341 agttgatcgg cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg cagggccaga
5401 ctggaggtgg caacgccaat cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg
5461 ttgggaatgt aattcagctc cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa
5521 acgtggctgg cctggttcac cacgcgggaa acggtctgat aagagacacc ggcatactct
5581 gcgacatcgt ataacgttac tggtttcaca ttcaccaccc tgaattgact ctcttccggg
5641 cgctatcatg ccataccgcg aaaggttttg cgccattcga tggtgtccgg gatctcgacg
5701 ctctccctta tgcgactcct gcattaggaa gcagcccagt agtaggttga ggccgttgag
5761 caccgccgcc gcaaggaatg gtgcatgcaa ggagatggcg cccaacagtc ccccggccac
5821 ggggcctgcc accataccca cgccgaaaca agcgctcatg agcccgaagt ggcgagcccg
5881 atcttcccca tcggtgatgt cggcgatata ggcgccagca accgcacctg tggcgccggt
5941 gatgccggcc acgatgcgtc cggcgtagag gatcgagatc gatctcgatc ccgcgaaat
//
