| 出品公司: | SIGMA |
|---|---|
| 载体名称: | p3XFLAG-CMV-8 |
| 质粒类型: | 哺乳动物表达载体； 分泌表达载体 |
| 高拷贝/低拷贝: | 低拷贝 |
| 克隆方法: | 多克隆位点；限制性内切酶 |
| 启动子: | CMV |
| 载体大小: | 4763 bp |
| 5' 测序引物及序列: | CMV Forward：CGCAAATGGGCGGTAGGCGTG |
| 3' 测序引物及序列: | -- |
| 载体标签: | 3XFLAG tag (N-末端） |
| 载体抗性: | 氨苄青霉素 |
| 筛选标记: | 无 |
| 克隆菌株: | -- |
| 宿主细胞（系）: | 常用细胞系，如293、Hela等 |
| 备注: |
p3XFLAG-CMV-8载体N端含有信号肽PPT ，辅助目的蛋白分泌表达； CMV启动子驱动目的蛋白高水平表达； FLAG标签含8个氨基酸残基（DYKDDDDK），3XFLAG标签含22个氨基酸残基， 一般位于融合蛋白的外表面，具有高度免疫灵 敏性，便于检测和纯化目的蛋白； 用EK蛋白酶可将3XFLAG标签切除。 |
| 产品目录号: | E9908 |
| 稳定性: | 瞬表达 |
| 组成型/诱导型: | 组成型 |
| 病毒/非病毒: | 非病毒 |
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
FLAG 和 3xFLAG 标签简介
A Proven System for Detection and Purification of Proteins The FLAG Expression System is an established way to express, purify and detect recombinant fusion proteins. FLAG and 3xFLAG have proven utility in numerous applications such as Western blotting, immunocytochemistry, immunoprecipitation, flow cytometry, protein purification, and in the study of protein-protein interactions, cell ultrastructure and protein localization. These small hydrophilic tags facilitate superior detection and purification of recombinant fusion proteins when using our highly specific and sensitive ANTI-FLAG antibodies. Ideal Epitope Tags: Small, Hydrophilic and Cleavable The FLAG expression system utilizes a short, hydrophilic 8-amino acid peptide that is fused to the recombinant protein of interest. Because of its hydrophilic nature, the FLAG peptide is likely to be located on the surface of the fusion protein. As a result, the FLAG peptide is easily accessible for cleavage by enterokinase (Ek) and for detection with antibodies. In addition, because of the small size of the FLAG peptide tag, it is not likely to obscure other epitopes, domains, or alter function, secretion, or transport of the fusion protein. The 3xFLAG system is an improvement upon the original system by fusing 3 tandem FLAG epitopes (22 amino acids). Detection of fusion proteins containing 3xFLAG is enhanced up to 200 times more than any other system. Like the original FLAG tag, 3xFLAG is hydrophilic, contains an Ek cleavage site, and is relatively small. Therefore, the risk of altering protein function, blocking other epitopes or decreasing solubility is minimized.
3xFLAG System Expression Vectors for Ultra-Sensitive
重组蛋白的检测
The 3xFLAG system is an improvement upon the original system by fusing 3 tandem FLAG epitopes for a total of 22 amino acids. Detection of fusion proteins containing 3xFLAG is enhanced up to 200 times more than any other system. Like the original FLAG tag, 3xFLAG is hydrophilic, contains an Enterokinase cleavage site, and is relatively small. Therefore, the risk of altering protein function, blocking other epitopes or decreasing solubility is minimized. In cases of low-level expression, 3xFLAG is ideal. Our selection of CMV vectors provides options for transient or stable expression of 3xFLAG fusion proteins.
Ultrasensitive System - Detect < 10 femtomoles using ANTI-FLAG Antibodies; Ideal for cases of low-level expression in mammalian cells.
Superior Detection - 20-200x more sensitive than any other system.
Versatile - Enhanced detection for immunoprecipitation, Western blots and immunocytochemistry.
Wide Selection of Detection and Purification Products - ANTI-FLAG antibodies, resins, and affinity capture plates.
CMV启动子：
Our mammalian expression vectors contain the strong CMV promoter for high-level constitutive expression in mammalian cells. The strong human cytomegalovirus (CMV) promoter regulatory region drives constitutive protein expression levels as high as 1 mg/L in COS cells. For less potent cell lines, protein levels are typically ~0.1 mg/L. The presence of the SV40 replication origin will result in high levels of DNA replication in SV40 replication permissive COS cells. Vectors containing the preprotrypsin leader (PPT) sequence direct secretion of FLAG fusion proteins into the culture medium for purification using ANTI-FLAG antibodies, resins, and plates.
p3XFLAG-CMV-8 Expression Vector is a 4.8 kb derivative of pCMV5 used to establish expression of N-terminal 3XFLAG fusion proteins in mammalian cells. The vector encodes three adjacent FLAG epitopes (Asp-Tyr-Lys-Xaa-Xaa-Asp) upstream of the multiple cloning region. This results in increased detection sensitivity using ANTI-FLAG M2 antibody. The third FLAG epitope includes the enterokinase recognition sequence, allowing cleavage of the 3XFLAG peptide from the purified fusion protein. The preprotrypsin leader sequence precedes the 3XFLAG sequence.
p3XFLAG-CMV-8 Expression Vector is a shuttle vector for E. coli and mammalian cells. Efficiency of replication is optimal when using an SV40 T antigen-expressing host.
p3XFLAG-CMV-7-BAP Control Plasmid is a 6.2 kb derivative of pCMV5 used for transient intracellular expression of N-terminal 3XFLAG bacterial alkaline phosphatase fusion protein in mammalian cells. The vector encodes three adjacent FLAG epitopes (Asp-Tyr-Lys-Xaa-Xaa-Asp) upstream of the multiple cloning region. This results in increased detection sensitivity using ANTI-FLAG M2 antibody. The third FLAG epitope includes the enterokinase recognition sequence, allowing cleavage of the 3XFLAG peptide from the purified fusion protein.
载体序列
LOCUS p3xFLAG-CMV8 4763 bp DNA SYN
DEFINITION p3xFLAG-CMV8
ACCESSION
KEYWORDS
SOURCE
ORGANISM other sequences; artificial sequences; vectors.
FEATURES Location/Qualifiers
source 1..4763
/organism="p3xFLAG-CMV8"
/mol_type="other DNA"
misc_feature complement(12..155)
/label="lacZ_a"
misc_feature 126..148
/label="M13_pUC_fwd_primer"
promoter 141..157
/label="M13_forward20_primer"
promoter 319..895
/label="CMV_immearly_promoter"
misc_feature 398..685
/label="CAG_enhancer"
promoter 841..912
/label="CMV_promoter"
misc_feature 852..872
/label="CMV_fwd_primer"
misc_feature 896..915
/label="pCEP_fwd_primer"
misc_feature 1028..1042
/label="EK"
terminator 1114..1580
/label="hGH_PA_terminator"
misc_feature complement(1750..1770)
/label="pBABE_3_primer"
misc_feature complement(1756..1971)
/label="SV40_enhancer"
promoter 1768..2036
/label="SV40_promoter"
rep_origin 1935..2012
/label="SV40_origin"
misc_feature 1997..2016
/label="SV40pro_F_primer"
promoter complement(2123..2141)
/label="T7_promoter"
misc_feature complement(2168..2190)
/label="M13_pUC_rev_primer"
promoter complement(2204..2233)
/label="lac_promoter"
gene complement(3316..4176)
/label="Ampicillin"
/gene="Ampicillin"
CDS complement(3316..4176)
/label="ORF frame 3"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY
IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVE
YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL
DRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL
LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA
EIGASLIKHW*"
promoter complement(4218..4246)
/label="AmpR_promoter"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY
IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVE
YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL
DRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL
LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA
EIGASLIKHW*"
rep_origin 4325..4631
/label="f1_origin"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY
IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVE
YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL
DRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL
LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA
EIGASLIKHW*"
ORIGIN
1 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
61 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
121 gttttcccag tcacgacgtt gtaaaacgac ggccagtgcc aagctgatct atacattgaa
181 tcaatattgg caattagcca tattagtcat tggttatata gcataaatca atattggcta
241 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
301 aatatgaccg ccatgttgac attgattatt gactagttat taatagtaat caattacggg
361 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
421 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
481 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
541 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
601 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
661 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
721 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
781 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
841 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
901 tcgtttagtg aaccgtcaga attaattcac catgtctgca cttctgatcc tagctcttgt
961 tggagctgca gttgctgact acaaagacca tgacggtgat tataaagatc atgacatcga
1021 ttacaaggat gacgatgaca agcttgcggc cgcgaattca tcgatagatc tgatatcggt
1081 accagtcgac tctagaggat cccgggtggc atccctgtga cccctcccca gtgcctctcc
1141 tggccctgga agttgccact ccagtgccca ccagccttgt cctaataaaa ttaagttgca
1201 tcattttgtc tgactaggtg tccttctata atattatggg gtggaggggg gtggtatgga
1261 gcaaggggca agttgggaag acaacctgta gggcctgcgg ggtctattgg gaaccaagct
1321 ggagtgcagt ggcacaatct tggctcactg caatctccgc ctcctgggtt caagcgattc
1381 tcctgcctca gcctcccgag ttgttgggat tccaggcatg catgaccagg ctcagctaat
1441 ttttgttttt ttggtagaga cggggtttca ccatattggc caggctggtc tccaactcct
1501 aatctcaggt gatctaccca ccttggcctc ccaaattgct gggattacag gcgtgaacca
1561 ctgctccctt ccctgtcctt ctgattttaa aataactata ccagcaggag gacgtccaga
1621 cacagcatag gctacctggc catgcccaac cggtgggaca tttgagttgc ttgcttggca
1681 ctgtcctctc atgcgttggg tccactcagt agatgcctgt tgaattgggt acgcggccag
1741 cttggctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
1801 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
1861 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
1921 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
1981 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc
2041 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc ctcgaggaac
2101 tgaaaaacca gaaagttaat tccctatagt gagtcgtatt aaattcgtaa tcatgtcata
2161 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
2221 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
2281 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
2341 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
2401 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
2461 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
2521 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
2581 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
2641 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
2701 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
2761 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
2821 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
2881 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
2941 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
3001 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
3061 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
3121 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
3181 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
3241 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
3301 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
3361 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
3421 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
3481 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
3541 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
3601 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
3661 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
3721 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
3781 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
3841 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
3901 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
3961 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
4021 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
4081 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
4141 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
4201 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
4261 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg cgccctgtag
4321 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
4381 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
4441 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
4501 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
4561 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
4621 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
4681 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa
4741 caaaatatta acgcttacaa ttt
//
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