| 质粒名称: | pBad-sfGFP |
|---|---|
| 出品公司: | addgene |
| 目录编号: | 85482 |
| 存储实验室: | Ryan Mehl |
|
载体骨架基本信息 |
|
| 载体名称: | pBAD/Myc-His A |
| 载体出品公司: | Invitrogen |
| 空载体大小: | 4100bp |
| 完整质粒大小: | 4737bp |
| 载体类型: | 细菌细胞表达 |
| 载体抗性: | 氨苄霉素 |
| 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: | superfolder GFP/sfGFP |
| 物种来源: | Synthetic |
| 插入大小: | 744bp |
| 基因突变: | T2A, S13L, A92T, V104T, V114I, E161K, Y193K, F198Y, D202T, I203V, Y258F, A283V, K288T, N290Y relative to RpBphP2 |
| GenBank编号 | JN247409 |
| 启动子: | Arabinose |
| 标签/融合蛋白: | 6x His (C terminal on insert) |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | NcoI(未破坏) |
| 3’克隆位点: | XhoI(未破坏) |
| 5’测序引物 | pBad forward |
| 3’测序引物 | pBad reverse |
|
文献引用方法 |
|
| 材料和方法部分: |
pBad-sfGFP was a gift from Ryan Mehl (Addgene plasmid # 85482 ; http://n2t.net/addgene:85482 ; RRID:Addgene_85482)
|
| 参考文献部分: | Generating permissive site-specific unnatural aminoacyl-tRNA synthetases. Miyake-Stoner SJ, Refakis CA, Hammill JT, Lusic H, Hazen JL, Deiters A, Mehl RA. Biochemistry. 2010 Mar 2;49(8):1667-77. doi: 10.1021/bi901947r. 10.1021/bi901947r PubMed 20082521 |
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 pBad-sfGFP 4737 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Arabinose inducible E. coli codon optimized superfolder GFP with
C-terminal His6 tag.
ACCESSION .
VERSION .
KEYWORDS pBad-sfGFP
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4737)
AUTHORS Miyake-Stoner SJ, Refakis CA, Hammill JT, Lusic H, Hazen JL, Deiters
A, Mehl RA
TITLE Generating permissive site-specific unnatural aminoacyl-tRNA
synthetases.
JOURNAL Biochemistry. 2010 Mar 2;49(8):1667-77. doi: 10.1021/bi901947r.
PUBMED 20082521
REFERENCE 2 (bases 1 to 4737)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..4737
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 1..285
/gene="araBAD"
/label=araBAD promoter
/note="promoter of the L-arabinose operon of E. coli; the
araC regulatory gene is transcribed in the opposite
direction (Guzman et al., 1995)"
primer_bind 208..227
/label=pBAD Forward
/note="For vectors with E. coli araBAD promoter, forward
primer"
CDS 319..1035
/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
ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLL
EFVTAAGITHGMDELYK"
CDS 1042..1059
/codon_start=1
/product="6xHis affinity tag"
/label=6xHis
/translation="HHHHHH"
primer_bind complement(1138..1155)
/label=pBAD Reverse
/note="For vectors with E. coli araBAD promoter, reverse
primer"
terminator 1308..1394
/gene="Escherichia coli rrnB"
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
terminator 1486..1513
/label=rrnB T2 terminator
/note="transcription terminator T2 from the E. coli rrnB
gene"
promoter 1532..1623
/gene="bla"
/label=AmpR promoter
CDS 1624..2484
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
primer_bind complement(1842..1861)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 2655..3243
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 3144..3163
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 3397..3414
/label=L4440
/note="L4440 vector, forward primer"
misc_feature 3429..3569
/label=bom
/note="basis of mobility region from pBR322"
primer_bind complement(3495..3514)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 3655..3677
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
CDS complement(3833..4711)
/codon_start=1
/gene="araC"
/product="L-arabinose regulatory protein"
/label=araC
/translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
EKVNDVAVKLS"
ORIGIN
1 aagaaaccaa ttgtccatat tgcatcagac attgccgtca ctgcgtcttt tactggctct
61 tctcgctaac caaaccggta accccgctta ttaaaagcat tctgtaacaa agcgggacca
121 aagccatgac aaaaacgcgt aacaaaagtg tctataatca cggcagaaaa gtccacattg
181 attatttgca cggcgtcaca ctttgctatg ccatagcatt tttatccata agattagcgg
241 atcctacctg acgcttttta tcgcaactct ctactgtttc tccatacccg ttttttgggc
301 taacaggagg aattaaccat ggttagcaaa ggtgaagaac tgtttaccgg cgttgtgccg
361 attctggtgg aactggatgg tgatgtgaat ggccataaat ttagcgttcg tggcgaaggc
421 gaaggtgatg cgaccaacgg taaactgacc ctgaaattta tttgcaccac cggtaaactg
481 ccggttccgt ggccgaccct ggtgaccacc ctgacctatg gcgttcagtg ctttagccgc
541 tatccggatc atatgaaacg ccatgatttc tttaaaagcg cgatgccgga aggctatgtg
601 caggaacgta ccattagctt caaagatgat ggcacctata aaacccgtgc ggaagttaaa
661 tttgaaggcg ataccctggt gaaccgcatt gaactgaaag gtattgattt taaagaagat
721 ggcaacattc tgggtcataa actggaatat aatttcaaca gccataatgt gtatattacc
781 gccgataaac agaaaaatgg catcaaagcg aactttaaaa tccgtcacaa cgtggaagat
841 ggtagcgtgc agctggcgga tcattatcag cagaataccc cgattggtga tggcccggtg
901 ctgctgccgg ataatcatta tctgagcacc cagagcgttc tgagcaaaga tccgaatgaa
961 aaacgtgatc atatggtgct gctggaattt gttaccgccg cgggcattac ccacggtatg
1021 gatgaactgt ataaaggcag ccaccatcat catcaccatt aaagctcgag atctgcagct
1081 ggtaccatat gggaattcga agcttggctg ttttggcgga tgagagaaga ttttcagcct
1141 gatacagatt aaatcagaac gcagaagcgg tctgataaaa cagaatttgc ctggcggcag
1201 tagcgcggtg gtcccacctg accccatgcc gaactcagaa gtgaaacgcc gtagcgccga
1261 tggtagtgtg gggtctcccc atgcgagagt agggaactgc caggcatcaa ataaaacgaa
1321 aggctcagtc gaaagactgg gcctttcgtt ttatctgttg tttgtcggtg aacgctctcc
1381 tgagtaggac aaatccgccg ggagcggatt tgaacgttgc gaagcaacgg cccggagggt
1441 ggcgggcagg acgcccgcca taaactgcca ggcatcaaat taagcagaag gccatcctga
1501 cggatggcct ttttgcgttt ctacaaactc ttttgtttat ttttctaaat acattcaaat
1561 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
1621 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
1681 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
1741 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
1801 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
1861 tcccgtgttg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
1921 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
1981 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
2041 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
2101 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
2161 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
2221 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
2281 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
2341 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
2401 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
2461 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
2521 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
2581 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
2641 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
2701 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
2761 aaggtaactg gcttcagcag agcgcagata ccaaatactg tccttctagt gtagccgtag
2821 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
2881 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
2941 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
3001 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
3061 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
3121 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
3181 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
3241 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
3301 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
3361 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
3421 gaagagcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata
3481 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta tacactccgc
3541 tatcgctacg tgactgggtc atggctgcgc cccgacaccc gccaacaccc gctgacgcgc
3601 cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga
3661 gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag cagatcaatt
3721 cgcgcgcgaa ggcgaagcgg catgcataat gtgcctgtca aatggacgaa gcagggattc
3781 tgcaaaccct atgctactcc gtcaagccgt caattgtctg attcgttacc aattatgaca
3841 acttgacggc tacatcattc actttttctt cacaaccggc acggaactcg ctcgggctgg
3901 ccccggtgca ttttttaaat acccgcgaga aatagagttg atcgtcaaaa ccaacattgc
3961 gaccgacggt ggcgataggc atccgggtgg tgctcaaaag cagcttcgcc tggctgatac
4021 gttggtcctc gcgccagctt aagacgctaa tccctaactg ctggcggaaa agatgtgaca
4081 gacgcgacgg cgacaagcaa acatgctgtg cgacgctggc gatatcaaaa ttgctgtctg
4141 ccaggtgatc gctgatgtac tgacaagcct cgcgtacccg attatccatc ggtggatgga
4201 gcgactcgtt aatcgcttcc atgcgccgca gtaacaattg ctcaagcaga tttatcgcca
4261 gcagctccga atagcgccct tccccttgcc cggcgttaat gatttgccca aacaggtcgc
4321 tgaaatgcgg ctggtgcgct tcatccgggc gaaagaaccc cgtattggca aatattgacg
4381 gccagttaag ccattcatgc cagtaggcgc gcggacgaaa gtaaacccac tggtgatacc
4441 attcgcgagc ctccggatga cgaccgtagt gatgaatctc tcctggcggg aacagcaaaa
4501 tatcacccgg tcggcaaaca aattctcgtc cctgattttt caccaccccc tgaccgcgaa
4561 tggtgagatt gagaatataa cctttcattc ccagcggtcg gtcgataaaa aaatcgagat
4621 aaccgttggc ctcaatcggc gttaaacccg ccaccagatg ggcattaaac gagtatcccg
4681 gcagcagggg atcattttgc gcttcagcca tacttttcat actcccgcca ttcagag
//
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