| 质粒名称: | pMK231 (AAVS1 CMV-MCS-PURO) |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 105924 |
| 存储实验室: | Masato Kanemaki |
|
载体骨架基本信息 |
|
| 载体名称: | pBluescript |
| 载体出品公司: | -- |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 7873 bp |
| 完整质粒大小: | -- |
| 载体修饰: | -- |
| 载体类型: | 哺乳细胞表达,CRISPR |
| 筛选标记: | 嘌呤霉素 |
| 高拷贝/低拷贝: | 高拷贝 |
| 生长条件: | 37 ℃ |
|
插入基因信息 |
|
| 插入基因名称: | -- |
| 物种来源: | -- |
| 插入大小: | -- |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子 | CMV |
| 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 | -- |
| 3' 测序引物 | -- |
|
文献引用方法 |
|
| 材料和方法部分: | pMK231 (AAVS1 CMV-MCS-PURO) was a gift from Masato Kanemaki (Addgene plasmid # 105924 ; http://n2t.net/addgene:105924 ; RRID:Addgene_105924) |
| 参考文献部分: | Dynein-Dynactin-NuMA clusters generate cortical spindle-pulling forces as a multi-arm ensemble. Okumura M, Natsume T, Kanemaki MT, Kiyomitsu T. Elife. 2018 May 31;7. pii: 36559. doi: 10.7554/eLife.36559. 10.7554/eLife.36559 PubMed 29848445 |
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
质粒图谱.png)
载体简介
载体序列
LOCUS pMK231 (AAVS1 CMV-MCS-PURO) 7867 bp ds-DNA circular SYN 13-AUG-2018
DEFINITION Donor for integration at the human AAVS1 locus.
ACCESSION .
VERSION .
KEYWORDS pMK231 (AAVS1 CMV-MCS-PURO)
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7867)
AUTHORS Okumura M, Natsume T, Kanemaki MT, Kiyomitsu T
TITLE Dynein-Dynactin-NuMA clusters generate cortical spindle-pulling
forces as a multi-arm ensemble.
JOURNAL Elife. 2018 May 31;7. pii: 36559. doi: 10.7554/eLife.36559.
PUBMED 29848445
REFERENCE 2 (bases 1 to 7867)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 13, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..7867
/organism="synthetic DNA construct"
/mol_type="other DNA"
protein_bind 107..128
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 143..173
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 181..197
/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 186..208
/label=M13/pUC Reverse
/note="In lacZ gene"
primer_bind 205..221
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 205..221
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
promoter 242..260
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
misc_feature 295..1098
/label=HA-L
/note="left homology arm from the adeno-associated virus
integration site (AAVS1) within intron 1 of the human
PPP1R12C gene"
enhancer 1155..1458
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 1459..1662
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 1613..1633
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
misc_feature 1703..1759
/label=MCS
/note="multiple cloning site"
polyA_signal 1884..2005
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(1921..1940)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 1975..1994
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
promoter 2014..2514
/label=hPGK promoter
/note="human phosphoglycerate kinase 1 promoter"
primer_bind 2433..2451
/label=hPGK-F
/note="Human PGK promoter, forward primer"
CDS 2536..3135
/codon_start=1
/gene="pac from Streptomyces alboniger"
/product="puromycin N-acetyltransferase"
/label=PuroR
/note="confers resistance to puromycin"
/translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
primer_bind complement(2536..2555)
/label=Puro-R
/note="Puromycin resistance gene, reverse primer. Also
called puro-variant-R"
primer_bind 3032..3052
/label=Puro-F
/note="Puromycin resistance gene, forward primer"
polyA_signal 3142..3366
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
misc_feature 3369..4205
/label=HA-R
/note="right homology arm from the adeno-associated virus
integration site (AAVS1) within intron 1 of the human
PPP1R12C gene"
primer_bind complement(4239..4258)
/label=T7
/note="T7 promoter, forward primer"
promoter complement(4240..4258)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(4265..4282)
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind complement(4265..4281)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(4274..4296)
/label=M13/pUC Forward
/note="In lacZ gene"
CDS 4419..4721
/codon_start=1
/gene="ccdB"
/product="CcdB, a bacterial toxin that poisons DNA gyrase"
/label=ccdB
/note="Plasmids containing the ccdB gene cannot be
propagated in standard E. coli strains."
/translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
primer_bind 4600..4619
/label=ccdB-fwd
/note="ccdB gene, forward primer"
CDS 5070..5864
/codon_start=1
/gene="aph(3')-II (or nptII)"
/product="aminoglycoside phosphotransferase from Tn5"
/label=NeoR/KanR
/note="confers resistance to neomycin, kanamycin, and G418
(Geneticin(R))"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
primer_bind complement(5124..5143)
/label=Neo-R
/note="Neomycin resistance gene, reverse primer"
primer_bind 5734..5753
/label=Neo-F
/note="Neomycin resistance gene, forward primer"
CDS complement(6114..6974)
/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 6737..6756
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 7098..7686
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 7587..7606
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 7840..7857
/label=L4440
/note="L4440 vector, forward primer"
ORIGIN
1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagctca
241 gaattaaccc tcactaaagg gactagtcct gcaggtttaa acgaattcgc cctttgcttt
301 ctctgaccag cattctctcc cctgggcctg tgccgctttc tgtctgcagc ttgtggcctg
361 ggtcacctct acggctggcc cagatccttc cctgccgcct ccttcaggtt ccgtcttcct
421 ccactccctc ttccccttgc tctctgctgt gttgctgccc aaggatgctc tttccggagc
481 acttccttct cggcgctgca ccacgtgatg tcctctgagc ggatcctccc cgtgtctggg
541 tcctctccgg gcatctctcc tccctcaccc aaccccatgc cgtcttcact cgctgggttc
601 ccttttcctt ctccttctgg ggcctgtgcc atctctcgtt tcttaggatg gccttctccg
661 acggatgtct cccttgcgtc ccgcctcccc ttcttgtagg cctgcatcat caccgttttt
721 ctggacaacc ccaaagtacc ccgtctccct ggctttagcc acctctccat cctcttgctt
781 tctttgcctg gacaccccgt tctcctgtgg attcgggtca cctctcactc ctttcatttg
841 ggcagctccc ctacccccct tacctctcta gtctgtgcta gctcttccag ccccctgtca
901 tggcatcttc caggggtccg agagctcagc tagtcttctt cctccaaccc gggcccctat
961 gtccacttca ggacagcatg tttgctgcct ccagggatcc tgtgtccccg agctgggacc
1021 accttatatt cccagggccg gttaatgtgg ctctggttct gggtactttt atctgtcccc
1081 tccaccccac agtggggcaa ttaatagtaa tcaattacgg ggtcattagt tcatagccca
1141 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
1201 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact
1261 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa
1321 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
1381 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta
1441 gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg
1501 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg
1561 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg
1621 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctggtttagt gaaccgtcag
1681 atccgctagc gctaccggac tcagatctcg agctcaagct tcgaattctg cagtcgacgg
1741 taccgcgggc ccgggatcca ccggggccgc gactctagat cataatcagc cataccacat
1801 ttgtagaggt tttacttgct ttaaaaaacc tcccacacct ccccctgaac ctgaaacata
1861 aaatgaatgc aattgttgtt gttaacttgt ttattgcagc ttataatggt tacaaataaa
1921 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
1981 tgtccaaact catcaatgta tcttaagttg gggttgcgcc ttttccaagg cagccctggg
2041 tttgcgcagg gacgcggctg ctctgggcgt ggttccggga aacgcagcgg cgccgaccct
2101 gggtctcgca cattcttcac gtccgttcgc agcgtcaccc ggatcttcgc cgctaccctt
2161 gtgggccccc cggcgacgct tcctgctccg cccctaagtc gggaaggttc cttgcggttc
2221 gcggcgtgcc ggacgtgaca aacggaagcc gcacgtctca ctagtaccct cgcagacgga
2281 cagcgccagg gagcaatggc agcgcgccga ccgcgatggg ctgtggccaa tagcggctgc
2341 tcagcagggc gcgccgagag cagcggccgg gaaggggcgg tgcgggaggc ggggtgtggg
2401 gcggtagtgt gggccctgtt cctgcccgcg cggtgttccg cattctgcaa gcctccggag
2461 cgcacgtcgg cagtcggctc cctcgttgac cgaatcaccg acctctctcc ccagggggat
2521 ccaccggagc ttaccatgac cgagtacaag cccacggtgc gcctcgccac ccgcgacgac
2581 gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc cacgcgccac
2641 accgtcgatc cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg
2701 cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc
2761 tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg
2821 gccgagttga gcggttcccg gctggccgcg cagcaacaga tggaaggcct cctggcgccg
2881 caccggccca aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag
2941 ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg
3001 gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc ccttctacga gcggctcggc
3061 ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc
3121 aagcccggtg cctgaggtac cctgtgcctt ctagttgcca gccatctgtt gtttgcccct
3181 cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg
3241 aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc
3301 aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct
3361 ctatggttac tagggacagg attggtgaca gaaaagcccc atccttaggc ctcctccttc
3421 ctagtctcct gatattgggt ctaaccccca cctcctgtta ggcagattcc ttatctggtg
3481 acacaccccc atttcctgga gccatctctc tccttgccag aacctctaag gtttgcttac
3541 gatggagcca gagaggatcc tgggagggag agcttggcag ggggtgggag ggaagggggg
3601 gatgcgtgac ctgcccggtt ctcagtggcc accctgcgct accctctccc agaacctgag
3661 ctgctctgac gcggctgtct ggtgcgtttc actgatcctg gtgctgcagc ttccttacac
3721 ttcccaagag gagaagcagt ttggaaaaac aaaatcagaa taagttggtc ctgagttcta
3781 actttggctc ttcacctttc tagtccccaa tttatattgt tcctccgtgc gtcagtttta
3841 cctgtgagat aaggccagta gccagccccg tcctggcagg gctgtggtga ggaggggggt
3901 gtccgtgtgg aaaactccct ttgtgagaat ggtgcgtcct aggtgttcac caggtcgtgg
3961 ccgcctctac tccctttctc tttctccatc cttctttcct taaagagtcc ccagtgctat
4021 ctgggacata ttcctccgcc cagagcaggg tcccgcttcc ctaaggccct gctctgggct
4081 tctgggtttg agtccttggc aagcccagga gaggcgctca ggcttccctg tcccccttcc
4141 tcgtccacca tctcatgccc ctggctctcc tgccccttcc ctacaggggt tcctggctct
4201 gctctaaggg cgaattcgcg gccgctaaat tcaattcgcc ctatagtgag tcgtattaca
4261 attcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta
4321 atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg
4381 atcgcccttc ccaacagttg cgcagcctat acgtacggca gtttaaggtt tacacctata
4441 aaagagagag ccgttatcgt ctgtttgtgg atgtacagag tgatattatt gacacgccgg
4501 ggcgacggat ggtgatcccc ctggccagtg cacgtctgct gtcagataaa gtctcccgtg
4561 aactttaccc ggtggtgcat atcggggatg aaagctggcg catgatgacc accgatatgg
4621 ccagtgtgcc ggtctccgtt atcggggaag aagtggctga tctcagccac cgcgaaaatg
4681 acatcaaaaa cgccattaac ctgatgttct ggggaatata aatgtcaggc atgagattat
4741 caaaaaggat cttcacctag atccttttca cgtagaaagc cagtccgcag aaacggtgct
4801 gaccccggat gaatgtcagc tactgggcta tctggacaag ggaaaacgca agcgcaaaga
4861 gaaagcaggt agcttgcagt gggcttacat ggcgatagct agactgggcg gttttatgga
4921 cagcaagcga accggaattg ccagctgggg cgccctctgg taaggttggg aagccctgca
4981 aagtaaactg gatggctttc ttgccgccaa ggatctgatg gcgcagggga tcaagctctg
5041 atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt
5101 ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct
5161 gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga
5221 ccgacctgtc cggtgccctg aatgaactgc aagacgaggc agcgcggcta tcgtggctgg
5281 ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact
5341 ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg
5401 agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct
5461 gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg
5521 gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt
5581 tcgccaggct caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg
5641 cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc
5701 ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag
5761 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt
5821 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgaattatt aacgcttaca
5881 atttcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatcaggt
5941 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
6001 aatatgtatc cgctcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
6061 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
6121 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
6181 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
6241 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
6301 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
6361 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
6421 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
6481 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
6541 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
6601 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
6661 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
6721 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
6781 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
6841 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
6901 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
6961 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
7021 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
7081 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
7141 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
7201 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
7261 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
7321 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
7381 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
7441 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
7501 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
7561 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
7621 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
7681 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
7741 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
7801 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
7861 gcggaag
//
