| 质粒名称: | pMK1200 |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 84219 |
| 存储实验室: | Martin Kampmann, Jonathan Weissman |
|
载体骨架基本信息 |
|
| 载体名称: | pSicoR |
| 载体出品公司: | -- |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | -- |
| 完整质粒大小: | -- |
| 载体修饰: | -- |
| 载体类型: | 哺乳动物表达，慢病毒，RNAi |
| 筛选标记: | 嘌呤霉素 |
| 高拷贝/低拷贝: | 未知 |
| 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 | -- |
| 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: | pMK1200 was a gift from Martin Kampmann & Jonathan Weissman (Addgene plasmid # 84219 ; http://n2t.net/addgene:84219 ; RRID:Addgene_84219) |
| 参考文献部分: | Integrated platform for genome-wide screening and construction of high-density genetic interaction maps in mammalian cells. Kampmann M, Bassik MC, Weissman JS. Proc Natl Acad Sci U S A. 2013 Jun 18;110(25):E2317-26. doi: 10.1073/pnas.1307002110. Epub 2013 Jun 5. 10.1073/pnas.1307002110 PubMed 23739767 |
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
LOCUS pMK1200 8626 bp ds-DNA circular SYN 06-AUG-2018
DEFINITION Lentiviral expression of shRNAs, compatible with double-shRNA
expression, EF1alpha promoter, mCherry.
ACCESSION .
VERSION .
KEYWORDS pMK1200
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 8626)
AUTHORS Kampmann M, Bassik MC, Weissman JS
TITLE Integrated platform for genome-wide screening and construction of
high-density genetic interaction maps in mammalian cells.
JOURNAL Proc Natl Acad Sci U S A. 2013 Jun 18;110(25):E2317-26. doi:
10.1073/pnas.1307002110. Epub 2013 Jun 5.
PUBMED 23739767
REFERENCE 2 (bases 1 to 8626)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 6, 2018 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..8626
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind complement(56..75)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 247..626
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 628..826
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
primer_bind 781..801
/label=CMV-F
/note="Human CMV immediate early promoter, forward primer"
LTR 844..1024
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 1071..1196
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 1695..1928
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
misc_feature 2455..2572
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
promoter 2675..2886
/label=EF-1-alpha core promoter
/note="core promoter for human elongation factor
EF-1-alpha"
intron 2887..3825
/label=EF-1-alpha intron A
/note="intron upstream of the start codon of human
EF-1-alpha"
primer_bind 3782..3802
/label=EF1a-F
/note="Human elongation factor-1a promoter, forward primer"
CDS 3846..4442
/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(3846..3865)
/label=Puro-R
/note="Puromycin resistance gene, reverse primer. Also
called puro-variant-R"
primer_bind 4342..4362
/label=Puro-F
/note="Puromycin resistance gene, forward primer"
CDS 4452..4505
/codon_start=1
/product="2A peptide from Thosea asigna virus capsid
protein"
/label=T2A
/note="Eukaryotic ribosomes fail to insert a peptide bond
between the Gly and Pro residues, yielding separate
polypeptides."
/translation="EGRGSLLTCGDVEENPGP"
regulatory 4512..4521
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 4518..5228
/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(4663..4681)
/label=mCherry-R
/note="mCherry, reverse primer"
primer_bind 4931..4950
/label=mCherry-F
/note="mCherry, forward primer"
ncRNA 5312..5326
/label=miR-30a loop
/note="loop from the the 71-nt precursor of the human
miR-30a microRNA (Zeng et al., 2002)"
/ncRNA_class="miRNA"
misc_feature 5515..6103
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(5568..5588)
/label=WPRE-R
/note="WPRE, reverse primer"
CDS complement(5986..5997)
/codon_start=1
/product="Factor Xa recognition and cleavage site"
/label=Factor Xa site
/translation="IEGR"
primer_bind complement(6106..6122)
/label=KS primer
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(6107..6123)
/label=pBluescriptKS
/note="For pBluescript vector"
LTR 6632..6812
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
rep_origin complement(6874..7462)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(6954..6973)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(7633..8493)
/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 8256..8275
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(8494..8598)
/gene="bla"
/label=AmpR promoter
ORIGIN
1 gcttaagcgg tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa
61 tctgctctga tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg
121 ctgagtagtg cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca
181 tgaagaatct gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata
241 cgcgttgaca ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc
301 atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac
361 cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa
421 tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag
481 tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc
541 ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct
601 acgtattagt catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg
661 gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt
721 tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga
781 cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagcgcg ttttgcctgt
841 actgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta actagggaac
901 ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg tgcccgtctg
961 ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg gaaaatctct
1021 agcagtggcg cccgaacagg gacttgaaag cgaaagggaa accagaggag ctctctcgac
1081 gcaggactcg gcttgctgaa gcgcgcacgg caagaggcga ggggcggcga ctggtgagta
1141 cgccaaaaat tttgactagc ggaggctaga aggagagaga tgggtgcgag agcgtcagta
1201 ttaagcgggg gagaattaga tcgcgatggg aaaaaattcg gttaaggcca gggggaaaga
1261 aaaaatataa attaaaacat atagtatggg caagcaggga gctagaacga ttcgcagtta
1321 atcctggcct gttagaaaca tcagaaggct gtagacaaat actgggacag ctacaaccat
1381 cccttcagac aggatcagaa gaacttagat cattatataa tacagtagca accctctatt
1441 gtgtgcatca aaggatagag ataaaagaca ccaaggaagc tttagacaag atagaggaag
1501 agcaaaacaa aagtaagacc accgcacagc aagcggccgg ccgcgctgat cttcagacct
1561 ggaggaggag atatgaggga caattggaga agtgaattat ataaatataa agtagtaaaa
1621 attgaaccat taggagtagc acccaccaag gcaaagagaa gagtggtgca gagagaaaaa
1681 agagcagtgg gaataggagc tttgttcctt gggttcttgg gagcagcagg aagcactatg
1741 ggcgcagcgt caatgacgct gacggtacag gccagacaat tattgtctgg tatagtgcag
1801 cagcagaaca atttgctgag ggctattgag gcgcaacagc atctgttgca actcacagtc
1861 tggggcatca agcagctcca ggcaagaatc ctggctgtgg aaagatacct aaaggatcaa
1921 cagctcctgg ggatttgggg ttgctctgga aaactcattt gcaccactgc tgtgccttgg
1981 aatgctagtt ggagtaataa atctctggaa cagatttgga atcacacgac ctggatggag
2041 tgggacagag aaattaacaa ttacacaagc ttaatacact ccttaattga agaatcgcaa
2101 aaccagcaag aaaagaatga acaagaatta ttggaattag ataaatgggc aagtttgtgg
2161 aattggttta acataacaaa ttggctgtgg tatataaaat tattcataat gatagtagga
2221 ggcttggtag gtttaagaat agtttttgct gtactttcta tagtgaatag agttaggcag
2281 ggatattcac cattatcgtt tcagacccac ctcccaaccc cgaggggacc cgacaggccc
2341 gaaggaatag aagaagaagg tggagagaga gacagagaca gatccattcg attagtgaac
2401 ggatcggcac tgcgtgcgcc aattctgcag acaaatggca gtattcatcc acaattttaa
2461 aagaaaaggg gggattgggg ggtacagtgc aggggaaaga atagtagaca taatagcaac
2521 agacatacaa actaaagaat tacaaaaaca aattacaaaa attcaaaatt ttcgggttta
2581 ttacagggac agcagagatc cagtttggtt agtaccgggc ccgctctagg gccgcgtgga
2641 taaccgtatt accgccatgc atgtgcccgt cagtgggcag agcgcacatc gcccacagtc
2701 cccgagaagt tggggggagg ggtcggcaat tgaaccggtg cctagagaag gtggcgcggg
2761 gtaaactggg aaagtgatgt cgtgtactgg ctccgccttt ttcccgaggg tgggggagaa
2821 ccgtatataa gtgcagtagt cgccgtgaac gttctttttc gcaacgggtt tgccgccaga
2881 acacaggtaa gtgccgtgtg tggttcccgc gggcctggcc tctttacggg ttatggccct
2941 tgcgtgcctt gaattacttc cacctggctg cagtacgtga ttcttgatcc cgagcttcgg
3001 gttggaagtg ggtgggagag ttcgaggcct tgcgcttaag gagccccttc gcctcgtgct
3061 tgagttgagg cctggcctgg gcgctggggc cgccgcgtgc gaatctggtg gcaccttcgc
3121 gcctgtctcg ctgctttcga taagtctcta gccatttaaa atttttgatg acctgctgcg
3181 acgctttttt tctggcaaga tagtcttgta aatgcgggcc aagatctgca cactggtatt
3241 tcggtttttg gggccgcggg cggcgacggg gcccgtgcgt cccagcgcac atgttcggcg
3301 aggcggggcc tgcgagcgcg gccaccgaga atcggacggg ggtagtctca agctggccgg
3361 cctgctctgg tgcctggcct cgcgccgccg tgtatcgccc cgccctgggc ggcaaggctg
3421 gcccggtcgg caccagttgc gtgagcggaa agatggccgc ttcccggccc tgctgcaggg
3481 agctcaaaat ggaggacgcg gcgctcggga gagcgggcgg gtgagtcacc cacacaaagg
3541 aaaagggcct ttccgtcctc agccgtcgct tcatgtgact ccacggagta ccgggcgccg
3601 tccaggcacc tcgattagtt ctcgagcttt tggagtacgt cgtctttagg ttggggggag
3661 gggttttatg cgatggagtt tccccacact gagtgggtgg agactgaagt taggccagct
3721 tggcacttga tgtaattctc cttggaattt gccctttttg agtttggatc ttggttcatt
3781 ctcaagcctc agacagtggt tcaaagtttt tttcttccat ttcaggtgtc gtgagctagc
3841 ccaccatgac cgagtacaag cccacggtgc gcctcgccac ccgcgacgac gtcccccggg
3901 ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc cacgcgccac accgtcgacc
3961 cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg cgcgtcgggc
4021 tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc tggaccacgc
4081 cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg gccgagttga
4141 gcggttcccg gctggccgcg cagcaacaga tggaaggcct cctggcgccg caccggccca
4201 aggagcccgc gtggttcctg gccaccgtcg gcgtctcgcc cgaccaccag ggcaagggtc
4261 tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg gtgcccgcct
4321 tcctggagac ctccgcgccc cgcaacctcc ccttctacga gcggctcggc ttcaccgtca
4381 ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc aagcccggtg
4441 ccggatcggg agagggcaga ggaagtctgc taacatgcgg tgacgtcgag gagaatcctg
4501 gcccaccggt cgccaccatg gtgagcaagg gcgaggagga taacatggcc atcatcaagg
4561 agttcatgcg cttcaaggtg cacatggagg gctccgtgaa cggccacgag ttcgagatcg
4621 agggcgaggg cgagggccgc ccctacgagg gcacccagac cgccaagctg aaggtgacca
4681 agggtggccc cctgcccttc gcctgggaca tcctgtcccc tcagttcatg tacggctcca
4741 aggcctacgt gaagcacccc gccgacatcc ccgactactt gaagctgtcc ttccccgagg
4801 gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg cgtggtgacc gtgacccagg
4861 actcctccct gcaggacggc gagttcatct acaaggtgaa gctgcgcggc accaacttcc
4921 cctccgacgg ccccgtaatg cagaagaaga ccatgggctg ggaggcctcc tccgagcgga
4981 tgtaccccga ggacggcgcc ctgaagggcg agatcaagca gaggctgaag ctgaaggacg
5041 gcggccacta cgacgctgag gtcaagacca cctacaaggc caagaagccc gtgcagctgc
5101 ccggcgccta caacgtcaac atcaagttgg acatcacctc ccacaacgag gactacacca
5161 tcgtggaaca gtacgaacgc gccgagggcc gccactccac cggcggcatg gacgagctgt
5221 acaagtaaga attgcctagg tattcgcctt gactcgagaa gaaggtatat tgctgttgac
5281 ccagagcgtt gggcaggtct gaagttcatt agtgaagcca cagatgtaat gaacttcagg
5341 acctgcccac tgccttgggc ctcgggcttc aaggggtcag tcacactctt tccctacacg
5401 acgctcttcc gatctatacc acatatctag agttgaaccg actagtaagc caggtcgaga
5461 catgtttaag ggttccggtt ccactaggta caattcgata tcaagcttat cgataatcaa
5521 cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt
5581 acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct
5641 ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc
5701 gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg
5761 ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc
5821 acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc
5881 actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct gctcgcctgt
5941 gttgccacct ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca
6001 gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt
6061 cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcatcgata ccgtcgacct
6121 cgatcgagac ctagaaaaac atggagcaat cacaagtagc aatacagcag ctaccaatgc
6181 tgattgtgcc tggctagaag cacaagagga ggaggaggtg ggttttccag tcacacctca
6241 ggtaccttta agaccaatga cttacaaggc agctgtagat cttagccact ttttaaaaga
6301 aaagggggga ctggaagggc taattcactc ccaacgaaga caagatatcc ttgatctgtg
6361 gatctaccac acacaaggct acttccctga ttggcagaac tacacaccag ggccagggat
6421 cagatatcca ctgacctttg gatggtgcta caagctagta ccagttgagc aagagaaggt
6481 agaagaagcc aatgaaggag agaacacccg cttgttacac cctgtgagcc tgcatgggat
6541 ggatgacccg gagagagaag tattagagtg gaggtttgac agccgcctag catttcatca
6601 catggcccga gagctgcatc cggactgtac tgggtctctc tggttagacc agatctgagc
6661 ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa gcttgccttg
6721 agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga gatccctcag
6781 acccttttag tcagtgtgga aaatctctag cagcatgtga gcaaaaggcc agcaaaaggc
6841 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
6901 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
6961 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
7021 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
7081 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
7141 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
7201 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
7261 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
7321 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
7381 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
7441 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
7501 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
7561 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
7621 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
7681 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
7741 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
7801 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
7861 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
7921 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
7981 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
8041 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
8101 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
8161 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
8221 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
8281 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
8341 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
8401 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
8461 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
8521 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
8581 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgac
//
载体序列
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