| 质粒名称: | pBLO 62.5 |
|---|---|
| 出品公司: | 爱迪基因 |
| 目录编号: | 123124 |
| 存储实验室: | Jennifer Doudna, Benjamin Oakes |
|
载体骨架基本信息 |
|
| 载体名称: | -- |
| 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: | 7897 bp |
| 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: | CasX protein |
| 物种来源: | Planctomycetes |
| 插入大小: | 2931 bp |
| 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: | pBLO 62.5 was a gift from Jennifer Doudna & Benjamin Oakes (Addgene plasmid # 123124 ; http://n2t.net/addgene:123124 ; RRID:Addgene_123124) |
| 参考文献部分: | CasX enzymes comprise a distinct family of RNA-guided genome editors. Liu JJ, Orlova N, Oakes BL, Ma E, Spinner HB, Baney KLM, Chuck J, Tan D, Knott GJ, Harrington LB, Al-Shayeb B, Wagner A, Brotzmann J, Staahl BT, Taylor KL, Desmarais J, Nogales E, Doudna JA. Nature. 2019 Feb 4. pii: 10.1038/s41586-019-0908-x. doi: 10.1038/s41586-019-0908-x. 10.1038/s41586-019-0908-x PubMed 30718774 |
买家导航
pBLO62.5载体基本信息
pBLO62.5载体图谱质粒图谱和多克隆位点信息
pBLO62.5载体简介
pBLO62.5载体序列
LOCUS pBLO 62.5 7896 bp ds-DNA circular SYN 28-MAR-2019
DEFINITION Human expression vector for planctomyces CasX and sgRNA.
ACCESSION .
VERSION .
KEYWORDS pBLO 62.5
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7896)
AUTHORS Liu JJ, Orlova N, Oakes BL, Ma E, Spinner HB, Baney KLM, Chuck J,
Tan D, Knott GJ, Harrington LB, Al-Shayeb B, Wagner A, Brotzmann J,
Staahl BT, Taylor KL, Desmarais J, Nogales E, Doudna JA
TITLE CasX enzymes comprise a distinct family of RNA-guided genome
editors.
JOURNAL Nature. 2019 Feb 4. pii: 10.1038/s41586-019-0908-x. doi:
10.1038/s41586-019-0908-x.
PUBMED 30718774
REFERENCE 2 (bases 1 to 7896)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Mar 28, 2019 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..7896
/organism="synthetic DNA construct"
/mol_type="other DNA"
rep_origin 1..589
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 490..509
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
promoter 651..891
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind 651..671
/label=hU6-F
/note="Human U6 promoter, forward primer"
primer_bind 822..841
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
enhancer 1042..1327
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer;
contains an 18-bp deletion relative to the standard CMV
enhancer"
promoter 1329..1606
/label=chicken beta-actin promoter
intron 1607..1834
/label=hybrid intron
/note="hybrid between chicken beta-actin (CBA) and minute
virus of mice (MMV) introns (Gray et al., 2011)"
regulatory 1846..1855
/regulatory_class="other"
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
CDS 1858..1878
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 4822..4842
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 4843..4890
/codon_start=1
/product="two tandem FLAG(R) epitope tags"
/label=2xFLAG
/translation="DYKDDDDKDYKDDDDK"
CDS 4900..4956
/codon_start=1
/product="2A peptide from porcine teschovirus-1
polyprotein"
/label=P2A
/note="Eukaryotic ribosomes fail to insert a peptide bond
between the Gly and Pro residues, yielding separate
polypeptides."
/translation="ATNFSLLKQAGDVEENPGP"
CDS 4957..5553
/codon_start=1
/gene="pac from Streptomyces alboniger"
/product="puromycin N-acetyltransferase"
/label=PuroR
/note="confers resistance to puromycin"
/translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
primer_bind 5450..5470
/label=Puro-F
/note="Puromycin resistance gene, forward primer"
primer_bind complement(5587..5604)
/label=BGH-rev
/note="Bovine growth hormone terminator, reverse primer.
Also called BGH reverse"
polyA_signal 5593..5800
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
repeat_region 5809..5949
/label=AAV2 ITR
/note="inverted terminal repeat of adeno-associated virus
serotype 2"
repeat_region 5809..5938
/label=AAV2 ITR
rep_origin 6024..6479
/direction=RIGHT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(6111..6130)
/label=F1ori-R
/note="F1 origin, reverse primer"
primer_bind 6321..6342
/label=F1ori-F
/note="F1 origin, forward primer"
primer_bind complement(6496..6515)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 6615..6637
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind complement(6675..6693)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 6761..6865
/gene="bla"
/label=AmpR promoter
CDS 6866..7726
/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(7084..7103)
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
ORIGIN
1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt gagggcctat
661 ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag ataattggaa
721 ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga aagtaataat
781 ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat atgcttaccg
841 taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga cgaaacacca
901 catctggcgc gtttattcca ttactttgga gccagtccca gcgactatgt cgtatggacg
961 aagcgcttat ttatcggaga gaaaccgata agtaaaacgc atcaaaggcg tgtccggcga
1021 gggcgagttt tttttggtac ccgttacata acttacggta aatggcccgc ctggctgacc
1081 gcccaacgac ccccgcccat tgacgtcaat agtaacgcca atagggactt tccattgacg
1141 tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat
1201 gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc attgtgccca
1261 gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat
1321 taccatggtc gaggtgagcc ccacgttctg cttcactctc cccatctccc ccccctcccc
1381 acccccaatt ttgtatttat ttatttttta attattttgt gcagcgatgg gggcgggggg
1441 gggggggggg cgcgcgccag gcggggcggg gsggggsgrg gggsggggsg gggsgrggcg
1501 gagaggtgcg gcggcagcca atcagagcgg cgcgctccga aagtttcctt ttatggcgag
1561 gcggcggcgg cggcggccct ataaaaagcg aagcgcgcgg cgggcgggag tcgctgcgcg
1621 ctgccttcgc cccgtgcccc gctccgccgc cgcctcgcgc cgcccgcccc ggctctgact
1681 gaccgcgtta ctcccacagg tgagcgggcg ggacggccct tctcctccgg gctgtaatta
1741 gctgagcaag aggtaagggt ttaagggatg gttggttggt ggggtattaa tgtttaatta
1801 cctggagcac ctgcctgaaa tcactttttt tcaggttgga ccggtgccac catggcccca
1861 aagaagaagc ggaaggtcag catgcaagag atcaagagaa tcaacaagat cagaaggaga
1921 ctggtcaagg acagcaacac aaagaaggcc ggcaagacag gccccatgaa aaccctgctc
1981 gtcagagtga tgacccctga cctgagagag cggctggaaa acctgagaaa gaagcccgag
2041 aacatccctc agcctatcag caacaccagc agggccaacc tgaacaagct gctgaccgac
2101 tacaccgaga tgaagaaagc catcctgcac gtgtactggg aagagttcca gaaagacccc
2161 gtgggcctga tgagcagagt tgctcagccc gctcctaaga acatcgacca gagaaagctg
2221 atccccgtga aggacggcaa cgagagactg acctctagcg gctttgcctg cagccagtgt
2281 tgccagcctc tgtacgtgta caagctggaa caagtgaacg acaagggcaa gccccacacc
2341 aactacttcg gcagatgcaa cgtgtccgag cacgagaggc tgatcctgct gtctcctcac
2401 aagcccgagg ccaacgatga gctggtcaca tacagcctgg gcaagttcgg acagagagcc
2461 ctggacttct acagcatcca cgtgaccagg gagagcaatc accctgtgaa gcccctggaa
2521 cagatcggcg gcaatagctg tgcctctgga cctgtgggaa aagccctgag cgacgcctgt
2581 atgggagccg tggcatcctt cctgaccaag taccaggaca tcatcctgga acaccagaaa
2641 gtgatcaaga agaacgagaa aagactggcc aacctcaagg atatcgccag cgctaacggc
2701 ctggcctttc ctaagatcac cctgcctcca cagcctcaca ccaaagaggg catcgaggcc
2761 tacaacaacg tggtggccca gatcgtgatt tgggtcaacc tgaatctgtg gcagaagctg
2821 aagatcggca gggacgaagc caagccactg cagagactga agggcttccc tagcttccct
2881 ctggtggaaa gacaggccaa tgaagtggat tggtgggaca tggtctgcaa cgtgaagaag
2941 ctgatcaacg agaagaaaga ggatggcaag gttttctggc agaacctggc cggctacaag
3001 agacaagaag ccctgctgcc ttacctgagc agcgaagagg accggaagaa gggcaagaag
3061 ttcgccagat accagttcgg cgacctgctg ctgcacctgg aaaagaagca cggcgaggac
3121 tggggcaaag tgtacgatga ggcctgggag agaatcgaca agaaggtgga aggcctgagc
3181 aagcacatta agctggaaga ggaaagaagg agcgaggacg cccaatctaa agccgctctg
3241 accgattggc tgagagccaa ggccagcttt gtgatcgagg gcctgaaaga ggccgacaag
3301 gacgagttct gcagatgcga gctgaagctg cagaagtggt acggcgatct gagaggcaag
3361 cccttcgcca ttgaggccga gaacagcatc ctggacatca gcggcttcag caagcagtac
3421 aactgcgcct tcatttggca gaaagacggc gtcaagaaac tgaacctgta cctgatcatc
3481 aattacttca aaggcggcaa gctgcggttc aagaagatca aacccgaggc cttcgaggct
3541 aacagattct acaccgtgat caacaaaaag tccggcgaga tcgtgcccat ggaagtgaac
3601 ttcaacttcg acgaccccaa cctgattatc ctgcctctgg ccttcggcaa gagacagggc
3661 agagagttca tctggaacga tctgctgagc ctggaaaccg gctctctgaa gctggccaat
3721 ggcagagtga tcgagaaaac cctgtacaac aggagaacca gacaggacga gcctgctctg
3781 tttgtggccc tgaccttcga gagaagagag gtgctggaca gcagcaacat caagcccatg
3841 aacctgatcg gcatcgaccg gggcgagaat atccctgctg tgatcgccct gacagaccct
3901 gaaggatgcc cactgagcag attcaaggac tccctgggca accctacaca catcctgaga
3961 atcggcgaga gctacaaaga gaagcagagg acaatccagg ccgccaaaga ggtggaacag
4021 agaagagccg gcggatactc taggaagtac gccagcaagg ccaagaatct ggccgacgac
4081 atggtccgaa acaccgccag agatctgctg tactacgccg tgacacagga cgccatgctg
4141 atcttcgaga atctgagcag aggcttcggc cggcagggca agagaacctt tatggccgag
4201 aggcagtaca ccagaatgga agattggctc acagctaaac tggcctacga gggactgccc
4261 agcaagacct acctgtccaa aacactggcc cagtatacct ccaagacctg cagcaattgc
4321 ggcttcacca tcaccagcgc cgactacgac agagtgctgg aaaagctcaa gaaaaccgcc
4381 accggctgga tgaccaccat caacggcaaa gagctgaagg ttgagggcca gatcacctac
4441 tacaacaggt acaagaggca gaacgtcgtg aaggatctga gcgtggaact ggacagactg
4501 agcgaagaga gcgtgaacaa cgacatcagc agctggacaa agggcagatc aggcgaggct
4561 ctgagcctgc tgaagaagag gtttagccac agacctgtgc aagagaagtt cgtgtgcctg
4621 aactgcggct tcgagacaca cgccgatgaa caggctgccc tgaacattgc cagaagctgg
4681 ctgttcctga gaagccaaga gtacaagaag taccagacca acaagaccac cggcaacacc
4741 gacaagaggg cctttgtgga aacctggcag agcttctaca gaaaaaagct gaaagaagtc
4801 tggaagcccg ccgtgactag tccaaaaaag aagagaaagg tagattacaa agatgacgat
4861 gacaaagact acaaggatga tgatgataag ggatccggcg caacaaactt ctctctgctg
4921 aaacaagccg gagatgtcga agagaatcct ggaccgaccg agtacaagcc cacggtgcgc
4981 ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac
5041 taccccgcca cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg
5101 caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac
5161 ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc
5221 gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg
5281 gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcgga
5341 gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag
5401 gcggccgagc gcgccggggt gcccgccttc ctggagacct ccgcgccccg caacctcccc
5461 ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc
5521 acctggtgca tgacccgcaa gcccggtgcc tgaacgcgtt aagaattcct agagctcgct
5581 gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc
5641 cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg
5701 catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca
5761 agggggagga ttgggaagag aatagcaggc atgctgggga gcggccgcag gaacccctag
5821 tgatggagtt ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa
5881 aggtcgcccg acgcccgggc tttgcccggg cggcctcagt gagcgagcga gcgcgcagct
5941 gcctgcaggg gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
6001 gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
6061 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ttagcgcccg ctcctttcgc
6121 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
6181 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgattt
6241 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
6301 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaactctat
6361 ctcgggctat tcttttgatt tataagggat tttgccgatt tcggtctatt ggttaaaaaa
6421 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt
6481 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc
6541 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
6601 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
6661 cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat
6721 ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
6781 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
6841 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
6901 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
6961 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
7021 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
7081 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
7141 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
7201 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
7261 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
7321 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
7381 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
7441 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
7501 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
7561 atctggagcc ggtgagcgtg gaagccgcgg tatcattgca gcactggggc cagatggtaa
7621 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
7681 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
7741 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
7801 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
7861 agcgtcagac cccgtagaaa agatcaaagg atcttc
//
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