| 质粒名称: | pCTCON2 Aga2P-HRP |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 73152 |
| 存储实验室: | Alice Ting |
|
载体骨架基本信息 |
|
| 载体名称: | pCTCON2 |
| 空载体大小: | 55791bp |
| 完整质粒大小: | 7232bp |
| 载体类型: | Yeast Expression |
| 载体抗性: | 氨苄霉素 |
| 生长温度: | 37 ℃ |
| 宿主菌: | DH5a |
| 拷贝数: | 未知 |
|
插入基因信息 |
|
| 插入基因名称: | Aga2P-HRP |
| 物种来源: | S. cerevisiae (budding yeast), Synthetic |
| 插入大小: | 1380bp |
| 启动子: | GAL1 |
| 标签/融合蛋白: |
HA tag
myc tag
|
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | NheI (未破坏) |
| 3’克隆位点: | BamHI(未破坏) |
| 5’测序引物 | gactacgctctgcaggagaacttg |
|
文献引用方法 |
|
| 材料和方法部分: | pCTCON2 Aga2P-HRP was a gift from Alice Ting (Addgene plasmid # 73152 ; http://n2t.net/addgene:73152 ; RRID:Addgene_73152) |
| 参考文献部分: | A split horseradish peroxidase for the detection of intercellular protein-protein interactions and sensitive visualization of synapses. Martell JD, Yamagata M, Deerinck TJ, Phan S, Kwa CG, Ellisman MH, Sanes JR, Ting AY. Nat Biotechnol. 2016 May 30. doi: 10.1038/nbt.3563. 10.1038/nbt.3563 PubMed 27240195 |
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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
载体简介
载体序列
LOCUS pCTCON2 Aga2P-HRP 7232 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION Full-length horseradish peroxidase (HRP) fused to the C-terminus of
the yeast mating protein Aga2P. Galactose-inducible expression..
ACCESSION .
VERSION .
KEYWORDS pCTCON2 Aga2P-HRP
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7232)
AUTHORS Martell JD, Yamagata M, Deerinck TJ, Phan S, Kwa CG, Ellisman MH,
Sanes JR, Ting AY
TITLE A split horseradish peroxidase for the detection of intercellular
protein-protein interactions and sensitive visualization of
synapses.
JOURNAL Nat Biotechnol. 2016 May 30. doi: 10.1038/nbt.3563.
PUBMED 27240195
REFERENCE 2 (bases 1 to 7232)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..7232
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind complement(10..28)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
misc_feature 70..573
/label=CEN/ARS
/note="S. cerevisiae CEN6 centromere fused to an
autonomously replicating sequence"
promoter 610..714
/gene="bla"
/label=AmpR promoter
CDS 715..1575
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFFHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
primer_bind complement(933..952)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 1746..2334
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 2488..2505
/label=L4440
/note="L4440 vector, forward primer"
promoter 2658..2688
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 2696..2712
/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 2701..2723
/label=M13/pUC Reverse
/note="In lacZ gene"
primer_bind 2720..2736
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 2720..2736
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
promoter 2757..2775
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
promoter 2940..3603
/gene="GAL1 and GAL10 genes of S. cerevisiae"
/label=GAL1,10 promoter
/note="divergent inducible promoter, regulated by Gal4"
protein_bind 3156..3273
/label=UAS
/bound_moiety="Gal4"
/note="upstream activating sequence mediating
Gal4-dependent induction"
primer_bind 3567..3590
/label=GAL1
/note="S. cerevisiae GAL1 promoter, forward primer"
CDS 3922..3933
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
CDS 3934..3960
/codon_start=1
/product="HA (human influenza hemagglutinin) epitope tag"
/label=HA
/translation="YPYDVPDYA"
CDS 3967..3987
/codon_start=1
/product="tobacco etch virus (TEV) protease recognition and
cleavage site"
/label=TEV site
/translation="ENLYFQG"
CDS 4990..5019
/codon_start=1
/product="Myc (human c-Myc proto-oncogene) epitope tag"
/label=Myc
/translation="EQKLISEEDL"
primer_bind complement(5352..5371)
/label=T7
/note="T7 promoter, forward primer"
promoter complement(5353..5371)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(5378..5395)
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind complement(5378..5394)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(5387..5409)
/label=M13/pUC Forward
/note="In lacZ gene"
rep_origin 5539..5994
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(5626..5645)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 5836..5857
/label=F1ori-F
/note="F1 origin, forward primer"
CDS complement(6091..6765)
/codon_start=1
/gene="S. cerevisiae TRP1"
/product="phosphoribosylanthranilate isomerase, required
for tryptophan biosynthesis"
/label=TRP1
/note="yeast auxotrophic marker"
/translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
KK"
promoter complement(6766..7046)
/gene="S. cerevisiae TRP1"
/label=TRP1 promoter
primer_bind complement(7063..7082)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 7182..7204
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
ORIGIN
1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
61 cttaggacgg atcgcttgcc tgtaacttac acgcgcctcg tatcttttaa tgatggaata
121 atttgggaat ttactctgtg tttatttatt tttatgtttt gtatttggat tttagaaagt
181 aaataaagaa ggtagaagag ttacggaatg aagaaaaaaa aataaacaaa ggtttaaaaa
241 atttcaacaa aaagcgtact ttacatatat atttattaga caagaaaagc agattaaata
301 gatatacatt cgattaacga taagtaaaat gtaaaatcac aggattttcg tgtgtggtct
361 tctacacaga caagatgaaa caattcggca ttaatacctg agagcaggaa gagcaagata
421 aaaggtagta tttgttggcg atccccctag agtcttttac atcttcggaa aacaaaaact
481 attttttctt taatttcttt ttttactttc tatttttaat ttatatattt atattaaaaa
541 atttaaatta taattatttt tatagcacgt gatgaaaagg acccaggtgg cacttttcgg
601 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
661 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
721 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
781 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
841 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
901 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
961 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
1021 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
1081 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
1141 ccgaaggagc taaccgcttt ttttcacaac atgggggatc atgtaactcg ccttgatcgt
1201 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
1261 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
1321 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
1381 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
1441 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
1501 ggcagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
1561 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
1621 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
1681 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
1741 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
1801 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
1861 ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac
1921 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
1981 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
2041 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
2101 acgacctaca ccgaactgag atacctacag cgtgagcatt gagaaagcgc cacgcttccc
2161 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
2221 agggagcttc caggggggaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
2281 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc cgagcctatg gaaaaacgcc
2341 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
2401 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc
2461 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc
2521 ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac
2581 aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag ttacctcact
2641 cattaggcac cccaggcttt acactttatg cttccggctc ctatgttgtg tggaattgtg
2701 agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa gctcggaatt
2761 aaccctcact aaagggaaca aaagctgggt acccgacagg ttatcagcaa caacacagtc
2821 atatccattc tcaattagct ctaccacagt gtgtgaacca atgtatccag caccacctgt
2881 aaccaaaaca attttagaag tactttcact ttgtaactga gctgtcattt atattgaatt
2941 ttcaaaaatt cttacttttt ttttggatgg acgcaaagaa gtttaataat catattacat
3001 ggcattacca ccatatacat atccatatac atatccatat ctaatcttac ttatatgttg
3061 tggaaatgta aagagcccca ttatcttagc ctaaaaaaac cttctctttg gaactttcag
3121 taatacgctt aactgctcat tgctatattg aagtacggat tagaagccgc cgagcgggtg
3181 acagccctcc gaaggaagac tctcctccgt gcgtcctcgt cttcaccggt cgcgttcctg
3241 aaacgcagat gtgcctcgcg ccgcactgct ccgaacaata aagattctac aatactagct
3301 tttatggtta tgaagaggaa aaattggcag taacctggcc ccacaaacct tcaaatgaac
3361 gaatcaaatt aacaaccata ggatgataat gcgattagtt ttttagcctt atttctgggg
3421 taattaatca gcgaagcgat gatttttgat ctattaacag atatataaat gcaaaaactg
3481 cataaccact ttactaatac tttcaacatt ttcggtttgt attacttctt attcaaatgt
3541 aataaaagta tcaacaaaaa attgttaata tacctctata ctttaacgtc aaggagaaaa
3601 aaccccggat cgaattccct acttcataca ttttcaatta agatgcagtt acttcgctgt
3661 ttttcaatat tttctgttat tgcttcagtt ttagcacagg aactgacaac tatatgcgag
3721 caaatcccct caccaacttt agaatcgacg ccgtactctt tgtcaacgac tactattttg
3781 gccaacggga aggcaatgca aggagttttt gaatattaca aatcagtaac gtttgtcagt
3841 aattgcggtt ctcacccctc aacaactagc aaaggcagcc ccataaacac acagtatgtt
3901 tttaaggaca atagctcgac gattgaaggt agatacccat acgacgttcc agactacgct
3961 ctgcaggaga acttgtactt tcaaggtctg caggctagtg gtggaggagg ctctggtgga
4021 ggcggtagcg gaggcggagg gtcggctagc cagcttactc caactttcta cgataactct
4081 tgtcctaacg tgagcaacat cgtgcgagat actatcgtga acgagttgcg gtctgatccg
4141 aggatagccg cttcaatttt gcggctgcat ttccacgatt gttttgtaaa cggctgtgat
4201 gcttccatcc tcctcgacaa taccacctct ttcaggactg aaaaggatgc cttcggtaac
4261 gcgaatagtg ctagagggtt tcccgtgatc gatagaatga aagccgctgt ggagagcgcc
4321 tgcccccgga cagttagttg tgcagacctt ctgaccatag cggcccagca atccgtgacg
4381 ctggccggtg gccccagctg gagggtcccc ctgggaagga gggactctct gcaggctttt
4441 ctggacttgg caaacgcaaa cctgcctgct cccttcttca cccttcccca gctgaaggat
4501 tccttcagga atgtcggcct gaaccgaagt agtgatttgg tggctctgtc cggaggccac
4561 acattcggca agaatcagtg caggttcatc atggacagac tgtataactt ttccaacacc
4621 ggcttgcctg accctaccct gaacacaaca tacttgcaga ctctcagagg gctgtgtccg
4681 ctcaatggca acttgtctgc actggttgac tttgacctgc gcaccccaac catcttcgac
4741 aataagtatt acgtcaatct ggaggaacag aagggcctga ttcagtccga ccaggagctg
4801 ttcagttccc caaatgctac cgacactatt cctcttgtgc gcagcttcgc taatagcaca
4861 caaacctttt tcaatgcttt cgtggaagct atggatcgga tgggaaatat aacgcccctg
4921 acgggtacgc agggacagat aagactgaac tgtagggtcg tgaacagcaa ctctggctcg
4981 ggcggatccg aacaaaagct tatttctgaa gaggacttgt aatagctcga gatctgataa
5041 caacagtgta gatgtaacaa aatcgacttt gttcccactg tacttttagc tcgtacaaaa
5101 tacaatatac ttttcatttc tccgtaaaca acatgttttc ccatgtaata tccttttcta
5161 tttttcgttc cgttaccaac tttacacata ctttatatag ctattcactt ctatacacta
5221 aaaaactaag acaattttaa ttttgctgcc tgccatattt caatttgtta taaattccta
5281 taatttatcc tattagtagc taaaaaaaga tgaatgtgaa tcgaatccta agagaattga
5341 gctccaattc gccctatagt gagtcgtatt acaattcact ggccgtcgtt ttacaacgtc
5401 gtgactggga aaaccctggc gttacccaac ttaatcgcct tgcagcacat ccccccttcg
5461 ccagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc
5521 tgaatggcga atggcgcgac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
5581 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
5641 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
5701 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
5761 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
5821 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
5881 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
5941 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgtttaca atttcctgat
6001 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcaggcaag tgcacaaaca
6061 atacttaaat aaatactact cagtaataac ctatttctta gcatttttga cgaaatttgc
6121 tattttgtta gagtctttta caccatttgt ctccacacct ccgcttacat caacaccaat
6181 aacgccattt aatctaagcg catcaccaac attttctggc gtcagtccac cagctaacat
6241 aaaatgtaag ctttcggggc tctcttgcct tccaacccag tcagaaatcg agttccaatc
6301 caaaagttca cctgtcccac ctgcttctga atcaaacaag ggaataaacg aatgaggttt
6361 ctgtgaagct gcactgagta gtatgttgca gtcttttgga aatacgagtc ttttaataac
6421 tggcaaaccg aggaactctt ggtattcttg ccacgactca tctccatgca gttggacgat
6481 atcaatgccg taatcattga ccagagccaa aacatcctcc ttaggttgat tacgaaacac
6541 gccaaccaag tatttcggag tgcctgaact atttttatat gcttttacaa gacttgaaat
6601 tttccttgca ataaccgggt caattgttct ctttctattg ggcacacata taatacccag
6661 caagtcagca tcggaatcta gagcacattc tgcggcctct gtgctctgca agccgcaaac
6721 tttcaccaat ggaccagaac tacctgtgaa attaataaca gacatactcc aagctgcctt
6781 tgtgtgctta atcacgtata ctcacgtgct caatagtcac caatgccctc cctcttggcc
6841 ctctcctttt cttttttcga ccgaattaat tcttaatcgg caaaaaaaga aaagctccgg
6901 atcaagattg tacgtaaggt gacaagctat ttttcaataa agaatatctt ccactactgc
6961 catctggcgt cataactgca aagtacacat atattacgat gctgtctatt aaatgcttcc
7021 tatattatat atatagtaat gtcgtttatg gtgcactctc agtacaatct gctctgatgc
7081 cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg
7141 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca
7201 gaggttttca ccgtcatcac cgaaacgcgc ga
//
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