| 质粒名称: | pLEVI(408)-ColE-iLight-msfGFP |
|---|---|
| 出品公司： | addgene |
| 目录编号: | 170268 |
| 存储实验室: | Vladislav Verkhusha |
|
载体骨架基本信息 |
|
| 载体名称: | pLEVI(408)-ColE |
| 载体出品公司: | Y. Yang lab (East China University of Science and Technology, China) |
| 空载体大小: | 5347 bp |
| 完整质粒大小: | 7702 bp |
| 载体修饰: | iLight-msfGFP was inserted C-terminally in frame with LexADBD. |
| 载体类型: | 细胞表达 |
| 筛选标记: | mCherry |
| 载体抗性: | 大观霉素，50 μg/mL |
| 生长温度: | 37 ℃ |
| 宿主菌: | DH5a |
| 拷贝数: | 低拷贝 |
|
插入基因信息 |
|
| 插入基因名称: | iiLight |
| 物种来源: | Synthetic Idiomarina sp. A28L |
| 插入大小: | 2355 bp |
| 基因突变: | I68F, H80Q, A86T, R90S S242C, E274K, R295H I360V, and L464V. |
| GenBank编号 | MW890755 |
| 启动子: | pJ23116 |
| 标签/融合蛋白: | msfGFP (C terminal on insert) |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | SacII(未破坏) |
| 3’克隆位点: | SacII (未破坏) |
| 5’测序引物 | atgaaagcgttaacggccaggcaac |
| 3’测序引物 | cgtcgccgtccagctcgaccag |
|
文献引用方法 |
|
| 材料和方法部分: | pLEVI(408)-ColE-iLight-msfGFP was a gift from Vladislav Verkhusha (Addgene plasmid # 170268 ; http://n2t.net/addgene:170268 ; RRID:Addgene_170268) |
| 参考文献部分: | Single-component near-infrared optogenetic systems for gene transcription regulation. Kaberniuk AA, Baloban M, Monakhov MV, Shcherbakova DM, Verkhusha VV. Nat Commun. 2021 Jun 23;12(1):3859. doi: 10.1038/s41467-021-24212-7. 10.1038/s41467-021-24212-7 PubMed 34162879 |
买家导航
pLEVI(408)-ColE-iLight-msfGFP载体pLEVI(408)-ColE-iLight-msfGFP质粒基本信息
pLEVI(408)-ColE-iLight-msfGFP质粒图谱pLEVI(408)-ColE-iLight-msfGFP载体图谱和pLEVI(408)-ColE-iLight-msfGFP载体序列pLEVI(408)-ColE-iLight-msfGFP质粒序列多克隆位点信息
-ColE-iLight-msfGFP170268质粒图谱.png)
pLEVI(408)-ColE-iLight-msfGFP质粒pLEVI(408)-ColE-iLight-msfGFP载体简介
pLEVI(408)-ColE-iLight-msfGFP质粒序列pLEVI(408)-ColE-iLight-msfGFP载体序列
LOCUS pLEVI(408)-ColE-iLight-msfGFP 7703 bp ds-DNA circular SYN 15-JUN-2021
DEFINITION Expresses LexA_DBD-iLight-msfGFP under J23116 promoter and mCherry
under ColE promoter with a LexA408 operator in bacteria..
ACCESSION .
VERSION .
KEYWORDS pLEVI(408)-ColE-iLight-msfGFP
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7703)
TITLE Single-component near-infrared optogenetic systems for gene
transcription regulation
REFERENCE 2 (bases 1 to 7703)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Jun 15, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..7703
/organism="synthetic DNA construct"
/mol_type="other DNA"
CDS 1958..2677
/codon_start=1
/product="GFP variant that folds robustly even when fused
to poorly folded proteins (Pedelacq et al., 2006)"
/label=superfolder GFP
/note="mammalian codon-optimized"
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL
KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDD
GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIK
ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
primer_bind complement(2003..2024)
/label=EGFP-N
/note="EGFP, reverse primer"
primer_bind 2611..2632
/label=EGFP-C
/note="EGFP, forward primer"
terminator 2887..2933
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
primer_bind complement(2956..2974)
/label=T7 Term
/note="T7 terminator, reverse primer"
terminator 2970..3017
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
CDS complement(3188..3979)
/codon_start=1
/gene="aadA"
/product="aminoglycoside adenylyltransferase (Murphy,
1985)"
/label=SmR
/note="confers resistance to spectinomycin and
streptomycin"
/translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
promoter complement(3980..4071)
/gene="bla"
/label=AmpR promoter
rep_origin complement(4119..4857)
/direction=LEFT
/label=CloDF13 ori
/note="Plasmids containing the CloDF13 (CDF) origin of
replication can be propagated in E. coli cells that contain
additional plasmids with compatible origins."
CDS complement(5067..6149)
/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 6111..6130
/label=LacI-R
/note="LacI, reverse primer"
promoter complement(6150..6227)
/gene="lacI"
/label=lacI promoter
terminator 6453..6499
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
CDS 6675..7385
/codon_start=1
/product="monomeric derivative of DsRed fluorescent protein
(Shaner et al., 2004)"
/label=mCherry
/note="mammalian codon-optimized"
/translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
EGRHSTGGMDELYK"
primer_bind complement(6820..6838)
/label=mCherry-R
/note="mCherry, reverse primer"
primer_bind 7088..7107
/label=mCherry-F
/note="mCherry, forward primer"
terminator 7592..7638
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
ORIGIN
1 cagctagctc agtcctaggg actatgctag caaacactat aacttcctag gtattattat
61 gaaagcgtta acggccaggc aacaagaggt gtttgatctc atccgtgatc acatcagcca
121 gacaggtatg ccgccgacgc gtgcggaaat cgcgcagcgt ttggggttcc gttccgcgtc
181 ctcggctgaa gaacatctga aggcgctggc acgcaaaggc gttattgaaa ttgtttccgg
241 cgcatcacgc gggattcgtc tgttgcagga agaggaagaa gggttgccgc tggtaggtcg
301 tgtggctgcc ggtgaaccgc ggagtgctgg tggggcagca gatctgggta gtgatgatat
361 cagcaaactg attgcagcat gtgatcaaga accgattcat attccgaatg caattcagcc
421 gtttggtgca atgctgattg ttgaaaaaga tacccagcag attgtttatg caagcgcaaa
481 tagcgcagaa tatttcagcg ttgcagataa caccattcat gaactgagcg attttaaaca
541 ggccaacatt aatagcctgc tgccggaaca actgattagc ggtctgacaa gcgcaattag
601 tgaaaatgaa ccgatttggg ttgaaaccga tcgtctgagc tttctgggtt ggcgtcatga
661 aaactattac atcattgaag tggaacgcta tcatgtgcag accagcaatt ggtttgaaat
721 tcagtttcag cgtgcctttc agaaactgcg taattgcaaa acccataacg atctgattaa
781 taccctgacc cgtctgattc aagaaatcag cggttatgat cgcgtgatga tctatcaatt
841 tgatccggaa tggaatggtc gtgttattgc agaaagcgtt cgtcagctgt ttaccagcat
901 gctgaatcat cattttccgg caagcgatat tccggcacag gcacgtgcaa tgtatagcat
961 taatccgatt cgtattatcc cggatgttaa tgcagaaccg cagccgctgc acatgattca
1021 taaaccgcaa aataccgaag cagttaatct gagctgcggt gttctgcgtg cagttagccc
1081 tctgcacatg cagtatctgc gtaattttgg tgttagcgca agcaccagca ttggcatttt
1141 taacgaagat aaactgtggg gtatcgttgc atgccatcat accaaaccgc gtgcaattgg
1201 tcgtcgtatt cgtcatctgc tggttcgtac cgttgaattt gcagcagaac gtctgtggct
1261 gattcatagc cgtaatgttg aacgttatat ggttaccgtt caggcagcac gtgaacagct
1321 gagcaccacc gcagatgata aacattcaag ccatgaaatc gtgattgaac atgcagcaga
1381 ttggtgtaaa ctgtttcgtt gtgatggtgt tggttatctg cgtggagaag aactgaccac
1441 ctatggtgaa acaccggatc agaccaccat taacaaactg gttgaatggc tggaagagaa
1501 cggtaaaaaa agcctgtttt ggcatagcca catgctgaaa gaagatgcac cgggtctgct
1561 gccggatggt agccgttttg caggtctgct ggcaattccg ctgaaaagtg atgcagacct
1621 gtttagctat ctgctgctgt ttcgtgtggc acagaatgaa gttcgtacct gggcgggtaa
1681 accggaaaaa ctgagcgttg aaaccagcac tggcaccatg gtgggtccgc gtaaaagttt
1741 tgaagcatgg caggatgaag ttagcggtaa aagccagccg tggcgtaccg cacagctgta
1801 tgcagcacgt gatattgcac gtgatctgct gattgtggca gatagcatgc agctgaatct
1861 gctgaatgat cagctggcag atgcaaatga aaatctggaa aaactggcca gctttgatga
1921 tctgaccgtc gactccggtg gtggttctgg tggtggaatg gtgagcaagg gcgaggagct
1981 gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt
2041 cagcgtgcgc ggcgagggcg agggcgatgc caccaacggc aagctgaccc tgaagttcat
2101 ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg
2161 cgtgcagtgc ttcagccgct accccgacca catgaagcgc cacgacttct tcaagtccgc
2221 catgcccgaa ggctacgtcc aggagcgcac catcagcttc aaggacgacg gcacctacaa
2281 gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg
2341 catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca acttcaacag
2401 ccacaacgtc tatatcaccg ccgacaagca gaagaacggc atcaaggcca acttcaagat
2461 ccgccacaac gtggaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc
2521 catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccaagct
2581 gagcaaagat cccaacgaga aacgcgatca catggtcctg ctggagttcg tgaccgccgc
2641 cgggatcact ctcggcatgg acgagctgta caagtgagaa ttccccctgt tttggcggat
2701 gagagaagat tttcagcctg atacagatta aatcagaacg cagaagcggt ctgataaaac
2761 agaatttgcc tggcggcagt agcgcggtgg tcccacctga ccccatgccg aactcagaag
2821 tgaaacgccg tagcgccgat ggtagtgtgg ggtctcccca tgcgagagta gggaactgcc
2881 aggcatcaaa taaaacgaaa ggctcagtcg aaagactggg cctttcgttt tatctgttgt
2941 ttgctcgagc tgccaccgct gagcaataac tagcataacc ccttggggcc tctaaacggg
3001 tcttgagggg ttttttgctg aaacctcagg catttgagaa gcacacggtc acactgcttc
3061 cggtagtcaa taaaccggta aaccagcaat agacataagc ggctatttaa cgaccctgcc
3121 ctgaaccgac gaccgggtca tcgtggccgg atcttgcggc ccctcggctt gaacgaattg
3181 ttagacatta tttgccgact accttggtga tctcgccttt cacgtagtgg acaaattctt
3241 ccaactgatc tgcgcgcgag gccaagcgat cttcttcttg tccaagataa gcctgtctag
3301 cttcaagtat gacgggctga tactgggccg gcaggcgctc cattgcccag tcggcagcga
3361 catccttcgg cgcgattttg ccggttactg cgctgtacca aatgcgggac aacgtaagca
3421 ctacatttcg ctcatcgcca gcccagtcgg gcggcgagtt ccatagcgtt aaggtttcat
3481 ttagcgcctc aaatagatcc tgttcaggaa ccggatcaaa gagttcctcc gccgctggac
3541 ctaccaaggc aacgctatgt tctcttgctt ttgtcagcaa gatagccaga tcaatgtcga
3601 tcgtggctgg ctcgaagata cctgcaagaa tgtcattgcg ctgccattct ccaaattgca
3661 gttcgcgctt agctggataa cgccacggaa tgatgtcgtc gtgcacaaca atggtgactt
3721 ctacagcgcg gagaatctcg ctctctccag gggaagccga agtttccaaa aggtcgttga
3781 tcaaagctcg ccgcgttgtt tcatcaagcc ttacggtcac cgtaaccagc aaatcaatat
3841 cactgtgtgg cttcaggccg ccatccactg cggagccgta caaatgtacg gccagcaacg
3901 tcggttcgag atggcgctcg atgacgccaa ctacctctga tagttgagtc gatacttcgg
3961 cgatcaccgc ttccctcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
4021 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa atagctagct
4081 cactcggtcg ctacgctccg ggcgtgagac tgcggcgggc gctgcggaca catacaaagt
4141 tacccacaga ttccgtggat aagcagggga ctaacatgtg aggcaaaaca gcagggccgc
4201 gccggtggcg tttttccata ggctccgccc tcctgccaga gttcacataa acagacgctt
4261 ttccggtgca tctgtgggag ccgtgaggct caaccatgaa tctgacagta cgggcgaaac
4321 ccgacaggac ttaaagatcc ccaccgtttc cggcgggtcg ctccctcttg cgctctcctg
4381 ttccgaccct gccgtttacc ggatacctgt tccgcctttc tcccttacgg gaagtgtggc
4441 gctttctcat agctcacaca ctggtatctc ggctcggtgt aggtcgttcg ctccaagctg
4501 ggctgtaagc aagaactccc cgttcagccc gactgctgcg ccttatccgg taactgttca
4561 cttgagtcca acccggaaaa gcacggtaaa acgccactgg cagcagccat tggtaactgg
4621 gagttcgcag aggatttgtt tagctaaaca cgcggttgct cttgaagtgt gcgccaaagt
4681 ccggctacac tggaaggaca gatttggttg ctgtgctctg cgaaagccag ttaccacggt
4741 taagcagttc cccaactgac ttaaccttcg atcaaaccac ctccccaggt ggttttttcg
4801 tttacagggc aaaagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
4861 tctactgaac cgctctagat ttcagtgcaa tttatctctt caaatgtagc acctgaagtc
4921 agccccatac gatataagtt gtaattctca tgttagtcat gccccgcgcc caccggaagg
4981 agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc taatgagtga
5041 gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
5101 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgcc
5161 agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt caccgcctgg
5221 ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt
5281 ttgatggtgg ttaacggcgg gatataacat gagctgtctt cggtatcgtc gtatcccact
5341 accgagatgt ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc
5401 gccatctgat cgttggcaac cagcatcgca gtgggaacga tgccctcatt cagcatttgc
5461 atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc tatcggctga
5521 atttgattgc gagtgagata tttatgccag ccagccagac gcagacgcgc cgagacagaa
5581 cttaatgggc ccgctaacag cgcgatttgc tggtgaccca atgcgaccag atgctccacg
5641 cccagtcgcg taccgtcttc atgggagaaa ataatactgt tgatgggtgt ctggtcagag
5701 acatcaagaa ataacgccgg aacattagtg caggcagctt ccacagcaat ggcatcctgg
5761 tcatccagcg gatagttaat gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc
5821 gccgctttac aggcttcgac gccgcttcgt tctaccatcg acaccaccac gctggcaccc
5881 agttgatcgg cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg cagggccaga
5941 ctggaggtgg caacgccaat cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg
6001 ttgggaatgt aattcagctc cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa
6061 acgtggctgg cctggttcac cacgcgggaa acggtctgat aagagacacc ggcatactct
6121 gcgacatcgt ataacgttac tggtttcaca ttcaccaccc tgaattgact ctcttccggg
6181 cgctatcatg ccataccgcg aaaggttttg cgccattcga tggtgtccgg gatctcgacg
6241 ctcggtaccc ccctgttttg gcggatgaga gaagattttc agcctgatac agattaaatc
6301 agaacgcaga agcggtctga taaaacagaa tttgcctggc ggcagtagcg cggtggtccc
6361 acctgacccc atgccgaact cagaagtgaa acgccgtagc gccgatggta gtgtggggtc
6421 tccccatgcg agagtaggga actgccaggc atcaaataaa acgaaaggct cagtcgaaag
6481 actgggcctt tcgttttatc tgttgtttgc gctagcggat cctgtttttt tgatcgtttt
6541 cacaaaaatg gaagtacaca gtcttgacag ggaaaatgca gcggcgtagc ttttatgccg
6601 tatataaaaa cggcgtttat atgtacggta tttattttta acttattgtt ttaaaagtca
6661 aagaggattt tataatggtg agcaagggcg aggaggataa catggccatc atcaaggagt
6721 tcatgcgctt caaggtgcac atggagggct ccgtgaacgg ccacgagttc gagatcgagg
6781 gcgagggcga gggccgcccc tacgagggca cccagaccgc caagctgaag gtgaccaagg
6841 gtggccccct gcccttcgcc tgggacatcc tgtcccctca gttcatgtac ggctccaagg
6901 cctacgtgaa gcaccccgcc gacatccccg actacttgaa gctgtccttc cccgagggct
6961 tcaagtggga gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg acccaggact
7021 cctccctgca ggacggcgag ttcatctaca aggtgaagct gcgcggcacc aacttcccct
7081 ccgacggccc cgtaatgcag aagaagacca tgggctggga ggcctcctcc gagcggatgt
7141 accccgagga cggcgccctg aagggcgaga tcaagcagag gctgaagctg aaggacggcg
7201 gccactacga cgctgaggtc aagaccacct acaaggccaa gaagcccgtg cagctgcccg
7261 gcgcctacaa cgtcaacatc aagttggaca tcacctccca caacgaggac tacaccatcg
7321 tggaacagta cgaacgcgcc gagggccgcc actccaccgg cggcatggac gagctgtaca
7381 agtaaagatc tctgttttgg cggatgagag aagattttca gcctgataca gattaaatca
7441 gaacgcagaa gcggtctgat aaaacagaat ttgcctggcg gcagtagcgc ggtggtccca
7501 cctgacccca tgccgaactc agaagtgaaa cgccgtagcg ccgatggtag tgtggggtct
7561 ccccatgcga gagtagggaa ctgccaggca tcaaataaaa cgaaaggctc agtcgaaaga
7621 ctgggccttt cgttttatct gttgtttgcg ctagcggatc cgcctatgca gcgacaaata
7681 ttgatagcct gaatcagtat tga
//
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pLEVI(408)-ColE-iLight-msfGFP质粒pLEVI(408)-ColE-iLight-msfGFP载体应用举例
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