| 质粒名称: | pLKO.1-scramble shRNA, Scramble shRNA |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 1864 |
| 质粒用途: | 第三代慢病毒质粒，表达scrambled shRNA的慢病毒质粒。 |
| 存储实验室: | David Sabatini |
|
载体骨架基本信息 |
|
| 载体名称: | pLKO.1 |
| 载体出品公司: | Stewart SA, RNA 2003 Apr; 9(4):493-501. |
| 载体抗性: | 氨苄霉素 |
| 空载体大小: | 7032 bp |
| 载体修饰: | 人细胞色素c氧化酶亚基VIII（human cytochrome C oxidase subunit VIII） |
| 载体类型: | 哺乳细胞表达，慢病毒表达，RNAi 表达质粒 |
| 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: | Scramble shRNA |
| 插入大小: | 60 bp |
| 插入序列: | CCTAAGGTTAAGTCGCCCTCGCTCGAGCGAGGGCGACTTAACCTTAGG |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | Age I(未破坏) |
| 3' 克隆位点: | EcoR I (未破坏) |
| 5' 测序引物 | LKO.1 5' |
|
文献引用方法 |
|
| 材料和方法部分: | scramble shRNA was a gift from David Sabatini (Addgene plasmid # 1864 ; http://n2t.net/addgene:1864 ; RRID:Addgene_1864) |
| 参考文献部分: | Phosphorylation and regulation of Akt/PKB by the rictor-mTOR complex. Sarbassov DD, Guertin DA, Ali SM, Sabatini DM. Science 2005 Feb 18;307(5712):1098-101. 10.1126/science.1106148 PubMed 15718470 |
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pLKO.1-Scramble shRNA载体质粒基本信息
pLKO.1-Scramble shRNA质粒图谱载体图谱和pLKO.1-Scramble shRNA载体序列质粒序列多克隆位点信息
pLKO.1-Scramble shRNA质粒载体简介
pLKO.1-Scramble shRNA质粒序列载体序列
LOCUS Scramble shRNA 7085 bp ds-DNA circular SYN 12-MAY-2021
DEFINITION 3rd gen lentiviral negative control vector pLKO.1-Scramble shRNA containing scrambled
shRNA.
ACCESSION .
VERSION .
KEYWORDS scramble shRNA
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7085)
AUTHORS Sarbassov DD, Guertin DA, Ali SM, Sabatini DM
TITLE Phosphorylation and regulation of Akt/PKB by the rictor-mTOR
complex.
JOURNAL Science 2005 Feb 18;307(5712):1098-101.
PUBMED 15718470
REFERENCE 2 (bases 1 to 7085)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58
http://www.biofeng.com
FEATURES Location/Qualifiers
source 1..7085
/organism="synthetic DNA construct"
/mol_type="other DNA"
primer_bind 66..84
/label=hPGK-F
/note="Human PGK promoter, forward primer"
CDS 169..768
/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(169..188)
/label=Puro-R
/note="Puromycin resistance gene, reverse primer. Also
called puro-variant-R"
primer_bind 665..685
/label=Puro-F
/note="Puromycin resistance gene, forward primer"
LTR 896..1129
/label=3' LTR (Delta-U3)
/note="self-inactivating 3' long terminal repeat (LTR) from
HIV-1"
polyA_signal 1201..1322
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(1238..1257)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 1292..1311
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
rep_origin 1362..1497
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 1424..1443
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
primer_bind complement(1517..1536)
/label=T7
/note="T7 promoter, forward primer"
promoter complement(1518..1536)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(1546..1563)
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind complement(1546..1562)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(1555..1577)
/label=M13/pUC Forward
/note="In lacZ gene"
rep_origin 1704..2159
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(1791..1810)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 2001..2022
/label=F1ori-F
/note="F1 origin, forward primer"
promoter 2185..2289
/gene="bla"
/label=AmpR promoter
CDS 2290..3150
/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 complement(2508..2527)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
rep_origin 3321..3909
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 3810..3829
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
primer_bind 4063..4080
/label=L4440
/note="L4440 vector, forward primer"
protein_bind 4197..4218
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 4233..4263
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 4271..4287
/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 4276..4298
/label=M13/pUC Reverse
/note="In lacZ gene"
primer_bind 4295..4311
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 4295..4311
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind 4330..4350
/label=T3
/note="T3 promoter, forward primer"
promoter 4332..4350
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
promoter 4378..4604
/label=RSV promoter
/note="Rous sarcoma virus enhancer/promoter"
LTR 4605..4785
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 4832..4957
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 5450..5683
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 5868..5912
/codon_start=1
/product="antigenic peptide corresponding to amino acids
655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik
et al., 2013)"
/label=gp41 peptide
/note="recognized by the 2H10 single-chain llama nanobody"
/translation="KNEQELLELDKWASL"
promoter 6210..6450
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind 6210..6230
/label=hU6-F
/note="Human U6 promoter, forward primer"
primer_bind 6381..6400
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
misc_feature 6556..6673
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
promoter 6722..147
/label=hPGK promoter
/note="human phosphoglycerate kinase 1 promoter"
ORIGIN
1 cgggaagggg cggtgcggga ggcggggtgt ggggcggtag tgtgggccct gttcctgccc
61 gcgcggtgtt ccgcattctg caagcctccg gagcgcacgt cggcagtcgg ctccctcgtt
121 gaccgaatca ccgacctctc tccccagggg gatccaccgg agcttaccat gaccgagtac
181 aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg caccctcgcc
241 gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg ccacatcgag
301 cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg
361 tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag cgtcgaagcg
421 ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc ccggctggcc
481 gcgcagcaac agatggaagg cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc
541 ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg
601 ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga gacctccgcg
661 ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg
721 cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctgacg cccgccccac
781 gacccgcagc gcccgaccga aaggagcgca cgaccccatg catcggtacc tttaagacca
841 atgacttaca aggcagctgt agatcttagc cactttttaa aagaaaaggg gggactggaa
901 gggctaattc actcccaacg aagacaagat ctgctttttg cttgtactgg gtctctctgg
961 ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct
1021 caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt
1081 aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag tagtagttca
1141 tgtcatctta ttattcagta tttataactt gcaaagaaat gaatatcaga gagtgagagg
1201 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
1261 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
1321 tatcatgtct ggctctagct atcccgcccc taactccgcc catcccgccc ctaactccgc
1381 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
1441 aggccgcctc ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
1501 ggacgtaccc aattcgccct atagtgagtc gtattacgcg cgctcactgg ccgtcgtttt
1561 acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc
1621 ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt
1681 gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
1741 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
1801 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
1861 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
1921 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
1981 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
2041 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
2101 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaattta
2161 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
2221 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
2281 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
2341 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
2401 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
2461 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
2521 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
2581 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
2641 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
2701 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
2761 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
2821 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
2881 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
2941 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
3001 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
3061 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
3121 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
3181 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
3241 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
3301 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
3361 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
3421 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
3481 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
3541 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
3601 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
3661 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
3721 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
3781 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
3841 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
3901 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
3961 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt
4021 gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag
4081 gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa
4141 tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat
4201 gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg
4261 ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac
4321 gccaagcgcg caattaaccc tcactaaagg gaacaaaagc tggagctgca agcttaatgt
4381 agtcttatgc aatactcttg tagtcttgca acatggtaac gatgagttag caacatgcct
4441 tacaaggaga gaaaaagcac cgtgcatgcc gattggtgga agtaaggtgg tacgatcgtg
4501 ccttattagg aaggcaacag acgggtctga catggattgg acgaaccact gaattgccgc
4561 attgcagaga tattgtattt aagtgcctag ctcgatacat aaacgggtct ctctggttag
4621 accagatctg agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat
4681 aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact
4741 agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc gcccgaacag
4801 ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc ggcttgctga
4861 agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa ttttgactag
4921 cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg ggagaattag
4981 atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata aattaaaaca
5041 tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc tgttagaaac
5101 atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga caggatcaga
5161 agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc aaaggataga
5221 gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca aaagtaagac
5281 caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg agggacaatt
5341 ggagaagtga attatataaa tataaagtag taaaaattga accattagga gtagcaccca
5401 ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata ggagctttgt
5461 tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg acgctgacgg
5521 tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg ctgagggcta
5581 ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag ctccaggcaa
5641 gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt tggggttgct
5701 ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt aataaatctc
5761 tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt aacaattaca
5821 caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag aatgaacaag
5881 aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata acaaattggc
5941 tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta agaatagttt
6001 ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta tcgtttcaga
6061 cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa gaaggtggag
6121 agagagacag agacagatcc attcgattag tgaacggatc tcgacggtat cgatcacgag
6181 actagcctcg agcggccgcc cccttcaccg agggcctatt tcccatgatt ccttcatatt
6241 tgcatatacg atacaaggct gttagagaga taattggaat taatttgact gtaaacacaa
6301 agatattagt acaaaatacg tgacgtagaa agtaataatt tcttgggtag tttgcagttt
6361 taaaattatg ttttaaaatg gactatcata tgcttaccgt aacttgaaag tatttcgatt
6421 tcttggcttt atatatcttg tggaaaggac gaaacaccgg tcctaaggtt aagtcgccct
6481 cgctcgagcg agggcgactt aaccttaggt ttttgaattc tcgacctcga gacaaatggc
6541 agtattcatc cacaatttta aaagaaaagg ggggattggg gggtacagtg caggggaaag
6601 aatagtagac ataatagcaa cagacataca aactaaagaa ttacaaaaac aaattacaaa
6661 aattcaaaat tttcgggttt attacaggga cagcagagat ccactttggc cgcggctcga
6721 gggggttggg gttgcgcctt ttccaaggca gccctgggtt tgcgcaggga cgcggctgct
6781 ctgggcgtgg ttccgggaaa cgcagcggcg ccgaccctgg gtctcgcaca ttcttcacgt
6841 ccgttcgcag cgtcacccgg atcttcgccg ctacccttgt gggccccccg gcgacgcttc
6901 ctgctccgcc cctaagtcgg gaaggttcct tgcggttcgc ggcgtgccgg acgtgacaaa
6961 cggaagccgc acgtctcact agtaccctcg cagacggaca gcgccaggga gcaatggcag
7021 cgcgccgacc gcgatgggct gtggccaata gcggctgctc agcagggcgc gccgagagca
7081 gcggc
//
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