| 质粒名称: | pBIS-GALkFLP(URA) |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 49460 |
| 存储实验室: | Marc Gartenberg |
|
载体骨架基本信息 |
|
| 载体名称: | pRS416 |
| 载体出品公司: | Stratagene |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 4898 bp |
| 完整质粒大小: | 6861 bp |
| 载体修饰: | -- |
| 载体类型: | 酵母表达 |
| 筛选标记: | URA3 |
| 高拷贝/低拷贝: | -- |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | Flp(H305L),Flp recombinase |
| 物种来源: | S. cerevisiae (budding yeast) |
| 插入大小: | -- |
| 基因突变: | H305L, ‘step-arrest’ mutant, |
| GenBank编号 | -- |
| 启动子 | GAL10 |
| 标签/融合蛋白: | -- |
| 克隆方法: | -- |
| 5' 克隆位点: | -- |
| 3' 克隆位点: | -- |
| 5' 测序引物 | M13-F20 |
| 3' 测序引物 | M13R |
|
文献引用方法 |
|
| 材料和方法部分: | pBIS-GALkFLP(URA) was a gift from Marc Gartenberg (Addgene plasmid # 49460 ; http://n2t.net/addgene:49460 ; RRID:Addgene_49460) |
| 参考文献部分: | Curing Saccharomyces cerevisiae of the 2 micron plasmid by targeted DNA damage. Tsalik EL, Gartenberg MR. Yeast. 1998 Jun 30;14(9):847-52. 10.1002/(SICI)1097-0061(19980630)14:9<847::AID-YEA285>3.0.CO;2-9 PubMed 9818722 |
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载体基本信息
载体图谱质粒图谱和多克隆位点信息
质粒图谱.png)
载体简介
载体序列
LOCUS pBIS-GALkFLP(URA) 6861 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION overespression of Flp step-arrest mutant in yeast, URA selection.
ACCESSION .
VERSION .
KEYWORDS pBIS-GALkFLP(URA)
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6861)
AUTHORS Tsalik EL, Gartenberg MR
TITLE Curing Saccharomyces cerevisiae of the 2 micron plasmid by targeted
DNA damage.
JOURNAL Yeast. 1998 Jun 30;14(9):847-52.
PUBMED 9818722
REFERENCE 2 (bases 1 to 6861)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..6861
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind complement(29..51)
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind 151..170
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
promoter 201..416
/gene="S. cerevisiae URA3"
/label=URA3 promoter
CDS 417..1220
/codon_start=1
/gene="S. cerevisiae URA3"
/product="orotidine-5'-phosphate decarboxylase, required
for uracil biosynthesis"
/label=URA3
/note="yeast auxotrophic marker, counterselectable with
5-fluoroorotic acid (5-FOA)"
/translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
rep_origin complement(1351..1806)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(1488..1509)
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind 1700..1719
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 1936..1958
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 1950..1967
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 1951..1967
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 1977..1996
/label=T7
/note="T7 promoter, forward primer"
promoter 1977..1995
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind 2020..2036
/label=pBluescriptKS
/note="For pBluescript vector"
primer_bind 2021..2037
/label=KS primer
/note="common sequencing primer, one of multiple similar
variants"
promoter 2250..2580
/gene="S. cerevisiae GAL10"
/label=GAL10 promoter
/note="inducible promoter, regulated by Gal4"
protein_bind 2250..2367
/label=UAS
/bound_moiety="Gal4"
/note="upstream activating sequence mediating
Gal4-dependent induction"
CDS 2588..3859
/codon_start=1
/product="site-specific recombinase"
/label=FLP
/note="FLP is a site-specific recombinase from
Saccharomyces cerevisiae. Recombination occurs at FRT
sequences."
/translation="MPQFGILCKTPPKVLVRQFVERFERPSGEKIALCAAELTYLCWMI
THNGTAIKRATFMSYNTIISNSLSFDIVNKSLQFKYKTQKATILEASLKKLIPAWEFTI
IPYYGQKHQSDITDIVSSLQLQFESSEEADKGNSHSKKMLKALLSEGESIWEITEKILN
SFEYTSRFTKTKTLYQFLFLATFINCGRFSDIKNVDPKSFKLVQNKYLGVIIQCLVTET
KTSVSRHIYFFSARGRIDPLVYLDEFLRNSEPVLKRVNRTGNSSSNKQEYQLLKDNLVR
SYNKALKKNAPYSIFAIKNGPKSLIGRHLMTSFLSMKGLTELTNVVGNWSDKRASAVAR
TTYTHQITAIPDHYFALVSRYYAYDPISKEMIALKDETNPIEEWQHIEQLKGSAEGSIR
YPAWNGIISQEVLDYLSSYINRRI"
primer_bind complement(4087..4107)
/label=T3
/note="T3 promoter, forward primer"
promoter complement(4087..4105)
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
primer_bind complement(4126..4142)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(4126..4142)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(4139..4161)
/label=M13/pUC Reverse
/note="In lacZ gene"
protein_bind 4150..4166
/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)."
promoter complement(4174..4204)
/label=lac promoter
/note="promoter for the E. coli lac operon"
primer_bind complement(4357..4374)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(4528..5116)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(4608..4627)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(5287..6147)
/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 5910..5929
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(6148..6252)
/gene="bla"
/label=AmpR promoter
misc_feature 6289..6792
/label=CEN/ARS
/note="S. cerevisiae CEN6 centromere fused to an
autonomously replicating sequence"
primer_bind 6834..6852
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
ORIGIN
1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
181 accataccac agcttttcaa ttcaattcat catttttttt ttattctttt ttttgatttc
241 ggtttctttg aaattttttt gattcggtaa tctccgaaca gaaggaagaa cgaaggaagg
301 agcacagact tagattggta tatatacgca tatgtagtgt tgaagaaaca tgaaattgcc
361 cagtattctt aacccaactg cacagaacaa aaacctgcag gaaacgaaga taaatcatgt
421 cgaaagctac atataaggaa cgtgctgcta ctcatcctag tcctgttgct gccaagctat
481 ttaatatcat gcacgaaaag caaacaaact tgtgtgcttc attggatgtt cgtaccacca
541 aggaattact ggagttagtt gaagcattag gtcccaaaat ttgtttacta aaaacacatg
601 tggatatctt gactgatttt tccatggagg gcacagttaa gccgctaaag gcattatccg
661 ccaagtacaa ttttttactc ttcgaagaca gaaaatttgc tgacattggt aatacagtca
721 aattgcagta ctctgcgggt gtatacagaa tagcagaatg ggcagacatt acgaatgcac
781 acggtgtggt gggcccaggt attgttagcg gtttgaagca ggcggcagaa gaagtaacaa
841 aggaacctag aggccttttg atgttagcag aattgtcatg caagggctcc ctatctactg
901 gagaatatac taagggtact gttgacattg cgaagagcga caaagatttt gttatcggct
961 ttattgctca aagagacatg ggtggaagag atgaaggtta cgattggttg attatgacac
1021 ccggtgtggg tttagatgac aagggagacg cattgggtca acagtataga accgtggatg
1081 atgtggtctc tacaggatct gacattatta ttgttggaag aggactattt gcaaagggaa
1141 gggatgctaa ggtagagggt gaacgttaca gaaaagcagg ctgggaagca tatttgagaa
1201 gatgcggcca gcaaaactaa aaaactgtat tataagtaaa tgcatgtata ctaaactcac
1261 aaattagagc ttcaatttaa ttatatcagt tattacccta tgcggtgtga aataccgcac
1321 agatgcgtaa ggagaaaata ccgcatcagg aaattgtaaa cgttaatatt ttgttaaaat
1381 tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa atcggcaaaa
1441 tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca gtttggaaca
1501 agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg
1561 gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg aggtgccgta
1621 aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg ggaaagccgg
1681 cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa
1741 gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg ccgctacagg
1801 gcgcgtcgcg ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg
1861 cctcttcgct attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg
1921 taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggccagtgag cgcgcgtaat
1981 acgactcact atagggcgaa ttgggtaccg ggccccccct cgaggtcgac ggtatcgata
2041 agcttgatat cgaattcctg cagcccgggg gatccaaaaa tcatcgcttc gctgattaat
2101 taccccagaa ataaggctaa aaaactaatc gcattatcat cctatggttg ttaatttgat
2161 tcgttcattt gaaggtttgt ggggccaggt tactgccaat ttttcctctt cataaccata
2221 aaagctagta ttgtagaatc tttattgttc ggagcagtgc ggcgcgaggc acatctgcgt
2281 ttcaggaacg cgaccggtga agacgaggac gcacggagga gagtcttcct tcggagggct
2341 gtcacccgct cggcggcttc taatccgtac ttcaatatag caatgagcag ttaagcgtat
2401 tactgaaagt tccaaagaga aggttttttt aggctaagat aatggggctc tttacatttc
2461 cacaacatat aagtaagatt agatatggat atgtatatgg atatgtatat ggtggtaatg
2521 ccatgtaata tgattattaa acttctttgc gtccatccaa aaaaaaagta agaatttttg
2581 ggtcgacatg ccacaatttg gtatattatg taaaacacca cctaaggtgc ttgttcgtca
2641 gtttgtggaa aggtttgaaa gaccttcagg tgagaaaata gcattatgtg ctgctgaact
2701 aacctattta tgttggatga ttacacataa cggaacagca atcaagagag ccacattcat
2761 gagctataat actatcataa gcaattcgct gagtttcgat attgtcaata aatcactcca
2821 gtttaaatac aagacgcaaa aagcaacaat tctggaagcc tcattaaaga aattgattcc
2881 tgcttgggaa tttacaatta ttccttacta tggacaaaaa catcaatctg atatcactga
2941 tattgtaagt agtttgcaat tacagttcga atcatcggaa gaagcagata agggaaatag
3001 ccacagtaaa aaaatgctta aagcacttct aagtgagggt gaaagcatct gggagatcac
3061 tgagaaaata ctaaattcgt ttgagtatac ttcgagattt acaaaaacaa aaactttata
3121 ccaattcctc ttcctagcta ctttcatcaa ttgtggaaga ttcagcgata ttaagaacgt
3181 tgatccgaaa tcatttaaat tagtccaaaa taagtatctg ggagtaataa tccagtgttt
3241 agtgacagag acaaagacaa gcgttagtag gcacatatac ttctttagcg caaggggtag
3301 gatcgatcca cttgtatatt tggatgaatt tttgaggaat tctgaaccag tcctaaaacg
3361 agtaaatagg accggcaatt cttcaagcaa taaacaggaa taccaattat taaaagataa
3421 cttagtcaga tcgtacaata aagctttgaa gaaaaatgcg ccttattcaa tctttgctat
3481 aaaaaatggc ccaaaatctc tcattggaag acatttgatg acctcatttc tttcaatgaa
3541 gggcctaacg gagttgacta atgttgtggg aaattggagc gataagcgtg cttctgccgt
3601 ggccaggaca acgtatactc atcagataac agcaatacct gatcactact tcgcactagt
3661 ttctcggtac tatgcatatg atccaatatc aaaggaaatg atagcattga aggatgagac
3721 taatccaatt gaggagtggc agcatataga acagctaaag ggtagtgctg aaggaagcat
3781 acgatacccc gcatggaatg ggataatatc acaggaggta ctagactacc tttcatccta
3841 cataaataga cgcatataag tacgcattta agcataaaca cgcactatgc cgttcttctc
3901 atgtatatat atatacaggc aacacgcaga tataggtgcg acgtgaacag tgagctgtat
3961 gtgcgcagct cgcgttgcat tttcggaagc gctcgttttc ggaaacgctt tgaagttcct
4021 attccgaagt tcctattctc tagttctaga gcggccgcca ccgcggtgga gctccagctt
4081 ttgttccctt tagtgagggt taattgcgcg cttggcgtaa tcatggtcat agctgtttcc
4141 tgtgtgaaat tgttatccgc tcacaattcc acacaacata ggagccggaa gcataaagtg
4201 taaagcctgg ggtgcctaat gagtgaggta actcacatta attgcgttgc gctcactgcc
4261 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
4321 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
4381 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
4441 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
4501 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
4561 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
4621 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
4681 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
4741 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
4801 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
4861 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
4921 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg
4981 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
5041 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
5101 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
5161 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
5221 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
5281 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
5341 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
5401 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
5461 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
5521 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
5581 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
5641 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
5701 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
5761 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
5821 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
5881 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
5941 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
6001 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
6061 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
6121 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
6181 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
6241 aggggttccg cgcacatttc cccgaaaagt gccacctggg tccttttcat cacgtgctat
6301 aaaaataatt ataatttaaa ttttttaata taaatatata aattaaaaat agaaagtaaa
6361 aaaagaaatt aaagaaaaaa tagtttttgt tttccgaaga tgtaaaagac tctaggggga
6421 tcgccaacaa atactacctt ttatcttgct cttcctgctc tcaggtatta atgccgaatt
6481 gtttcatctt gtctgtgtag aagaccacac acgaaaatcc tgtgatttta cattttactt
6541 atcgttaatc gaatgtatat ctatttaatc tgcttttctt gtctaataaa tatatatgta
6601 aagtacgctt tttgttgaaa ttttttaaac ctttgtttat ttttttttct tcattccgta
6661 actcttctac cttctttatt tactttctaa aatccaaata caaaacataa aaataaataa
6721 acacagagta aattcccaaa ttattccatc attaaaagat acgaggcgcg tgtaagttac
6781 aggcaagcga tccgtcctaa gaaaccatta ttatcatgac attaacctat aaaaataggc
6841 gtatcacgag gccctttcgt c
//
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