| 质粒名称: | pMaCTag-P34 |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 120030 |
| 存储实验室: | Michael Knop |
|
载体骨架基本信息 |
|
| 载体名称: | pFA6a |
| 载体出品公司: | -- |
| 载体抗性: | 氨苄青霉素 |
| 空载体大小: | 2412 bp |
| 完整质粒大小: | -- |
| 载体修饰: | -- |
| 载体类型: | PCR模板 |
| 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: | 3xFLAG |
| 物种来源: | H. sapiens (human) |
| 插入大小: | -- |
| 基因突变: | -- |
| GenBank编号 | -- |
| 启动子 | -- |
| 标签/融合蛋白: | 3xFLAG（主干上的C终端） |
| 克隆方法: | 限制性内切酶 |
| 5' 克隆位点: | BamHI(未破坏) |
| 3' 克隆位点: | SpeI (未破坏) |
| 5' 测序引物 | SP6 |
| 3' 测序引物 | AAACCACAACTAGAATGCAGTG |
|
文献引用方法 |
|
| 材料和方法部分: | pMaCTag-P34 was a gift from Michael Knop (Addgene plasmid # 120030 ; http://n2t.net/addgene:120030 ; RRID:Addgene_120030) |
| 参考文献部分: | CRISPR-Cas12a-assisted PCR tagging of mammalian genes. Fueller J, Herbst K, Meurer M, Gubicza K, Kurtulmus B, Knopf JD, Kirrmaier D, Buchmuller BC, Pereira G, Lemberg MK, Knop M. J Cell Biol. 2020 Jun 1;219(6). pii: 151766. doi: 10.1083/jcb.201910210. 10.1083/jcb.201910210 PubMed 32406907 |
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pMaCTag-P34载体 pMaCTagP34质粒基本信息
pMaCTag-P34质粒图谱 pMaCTagP34载体图谱和pMaCTag-P34载体序列 pMaCTagP34质粒序列多克隆位点信息
pMaCTag-P34质粒 pMaCTagP34载体简介
pMaCTag-P34质粒序列 pMaCTagP34载体序列
LOCUS pMaCTag-P34 4159 bp ds-DNA circular SYN 08-JAN-2019
DEFINITION Template for C-terminal PCR tagging of mammalian genes (Puromycin
selection) with 3xFLAG.
ACCESSION .
VERSION .
KEYWORDS pMaCTag-P34
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4159)
AUTHORS Fueller J, Meurer M, Herbst K, Gubicza K, Kurtulmus B, Knopf JD,
Kirrmaier D, Buchmuller B, Pereira G, Lemberg MK, Knop M
TITLE CRISPR/Cas12a-assisted PCR tagging of mammalian genes
JOURNAL bioRxiv
REFERENCE 2 (bases 1 to 4159)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Jan 8, 2019 from SnapGene Server 1.1.58
http://www.snapgene.com
FEATURES Location/Qualifiers
source 1..4159
/organism="synthetic DNA construct"
/mol_type="other DNA"
promoter 1..19
/label=SP6 promoter
/note="promoter for bacteriophage SP6 RNA polymerase"
primer_bind 1..18
/label=SP6
/note="SP6 promoter, forward primer"
CDS 81..146
/codon_start=1
/product="three tandem FLAG(R) epitope tags, followed by an
enterokinase cleavage site"
/label=3xFLAG
/translation="DYKDHDGDYKDHDIDYKDDDDK"
polyA_signal 156..277
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(193..212)
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
primer_bind 247..266
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
protein_bind 284..317
/label=loxP
/bound_moiety="Cre recombinase"
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (GCATACAT)."
promoter 324..640
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
rep_origin 491..626
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 553..572
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
polyA_signal 1262..1486
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
promoter 1493..1733
/label=U6 promoter
/note="RNA polymerase III promoter for human U6 snRNA"
primer_bind 1493..1513
/label=hU6-F
/note="Human U6 promoter, forward primer"
primer_bind 1664..1683
/label=LKO.1 5'
/note="Human U6 promoter, forward primer"
protein_bind 1743..1776
/label=loxP
/bound_moiety="Cre recombinase"
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (GCATACAT)."
primer_bind complement(1813..1832)
/label=T7
/note="T7 promoter, forward primer"
promoter complement(1814..1832)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(1919..1936)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(2090..2678)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(2170..2189)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(2849..3709)
/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 3472..3491
/label=Amp-R
/note="Ampicillin resistance gene, reverse primer"
promoter complement(3710..3814)
/gene="bla"
/label=AmpR promoter
primer_bind 3882..3900
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
primer_bind complement(3938..3960)
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind 4060..4079
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
ORIGIN
1 atttaggtga cactatagaa cgcggccgcc agctgaagct tcgtacgtca ggtggaggag
61 gtagtggcgg aggcggatcc gactacaaag accatgacgg tgattataaa gatcatgata
121 tcgattacaa ggatgacgat gacaagtaga ctagtaactt gtttattgca gcttataatg
181 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
241 ctagttgtgg tttgtccaaa ctcatcaatg tatcttagtc gatataactt cgtatagcat
301 acattatacg aagttatgtc gacggtgtgg aaagtcccca ggctccccag caggcagaag
361 tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc
421 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct
481 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg
541 actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa
601 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa gaattcgcca ccatgactga
661 gtacaagcct actgtgcgtt tagccaccag ggatgatgtg ccacgagccg tgaggacact
721 agcagccgcc tttgctgact acccagctac aaggcacacc gtggaccctg acagacacat
781 tgaaagagtc acagaacttc aggagctgtt tttaactagg gtgggcctgg atattggcaa
841 agtctgggtg gccgatgatg gagcagcagt tgcagtgtgg accaccccag agtctgttga
901 agctggagct gtctttgcag aaattggacc aagaatggcc gagttatctg gcagcaggct
961 ggctgctcaa cagcagatgg aaggattatt agctccccac agacccaagg agcctgcctg
1021 gttcctggca acagttggag tttctccaga ccatcagggc aagggcttag gatctgctgt
1081 ggtgctgcct ggtgtggaag cagctgagag agctggggtg ccagcatttc tggagacatc
1141 agctcctaga aacctgccct tctatgaacg gctgggcttc acagtgactg ctgatgtgga
1201 ggtgcctgag ggccccagaa cctggtgcat gacaagaaaa ccaggagcct gactgcagcg
1261 actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc
1321 ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt
1381 ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat
1441 tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggctcg aggagggcct
1501 atttcccatg attccttcat atttgcatat acgatacaag gctgttagag agataattgg
1561 aattaatttg actgtaaaca caaagatatt agtacaaaat acgtgacgta gaaagtaata
1621 atttcttggg tagtttgcag ttttaaaatt atgttttaaa atggactatc atatgcttac
1681 cgtaacttga aagtatttcg atttcttggc tttatatatc ttgtggaaag gacgaaacac
1741 cgataacttc gtatagcata cattatacga agttatggta ccgatgcagc tagcccgcgg
1801 atctgccggt ctccctatag tgagtcgtat taatttcgat aagccaggtt aacctgcatt
1861 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
1921 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
1981 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
2041 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
2101 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
2161 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
2221 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
2281 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
2341 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
2401 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
2461 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
2521 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
2581 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
2641 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
2701 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
2761 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
2821 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
2881 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
2941 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
3001 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
3061 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
3121 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
3181 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
3241 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
3301 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
3361 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
3421 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
3481 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
3541 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
3601 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
3661 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
3721 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
3781 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
3841 acgtctaaga aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc
3901 cctttcgtct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg
3961 agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt
4021 cagcgggtgt tggcgggtgt cggggctggc ttaactatgc ggcatcagag cagattgtac
4081 tgagagtgca ccatatggac atattgtcgt tagaacgcgg ctacaattaa tacataacct
4141 tatgtatcat acacatacg
//
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