pcDNA6/myc-His C

价格:1500元

联系方式:I47-825O-882O

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载体基本信息

出品公司: Invitrogen
载体名称: pcDNA6/myc-His C
质粒类型: 哺乳动物表达载体;cDNA表达载体
高拷贝/低拷贝: 高拷贝
克隆方法: 限制性内切酶,多克隆位点
启动子: CMV
载体大小: 5122 bp
5' 测序引物及序列: T7 Forward: 5’-TAATACGACTCACTATAGGG-3’
3' 测序引物及序列: BGH Reverse: 5’-TAGAAGGCACAGTCGAGG-3’
载体标签: myc Epitope(C-端);His tag(C-端)
载体抗性: 氨苄青霉素
筛选标记: Blasticidin
克隆菌株: TOP10F´, JM109,  INVaF´
宿主细胞(系): 常规细胞系,如293、Hela等
备注: pcDNA6/myc-His C载体是cDNA的表达与克隆载体;
CMV启动子驱动目的基因的过表达;
易于克隆到其它载体中去;
pcDNA6/myc-His A,B,C仅阅读框不同。
产品目录号: V221-20
稳定性: 瞬表达 或 稳表达
组成型/诱导型: 组成型
病毒/非病毒: 非病毒

载体质粒图谱和多克隆位点信息

pcDNA6-myc-His C载体图谱



pcDNA6-myc-His C 多克隆位点

pcDNA6-myc-His 载体特征1
pcDNA6-myc-His 载体特征2

载体简介

pcDNA.6/myc-His A, B, and C are 5.1-kb vectors designed for overproduction of recombinant proteins in mammalian cell lines. High-level stable and transient expression can be carried out in most mammalian cells. 

pcDNA.6/myc-His载体含有以下元件:

 Human cytomegalovirus immediate-early (CMV) promoter for high-level expression in a wide range of mammalian cells.
 Three reading frames to facilitate in-frame cloning with a C-terminal peptide encoding the myc (c-myc) epitope and a polyhistidine (6xHis) metal-binding tag.
 Blasticidin resistance gene (bsd) for selection of stable cell lines (Kimura et al.,1994).
Episomal replication in cell lines that are latently infected with SV40 or that express the SV40 large T antigen (e.g., COS7).
The control plasmid, pcDNA.6/myc-His/lacZ, is included for use as a positive control for transfection, expression, and detection in the cell line of choice.


实验流程

使用pcDNA6/myc-His 载体克隆和表达目的基因的实验流程如下:
 Consult the multiple cloning sites described on pages 3–5 to determine which vector (A, B, or C) should be used to clone your gene in-frame with the C-terminal myc epitope and the polyhistidine tag. Ligate your insert into the appropriate vector and transform into E. coli. Select transformants on 50–100 μg/ml ampicillin or 50 μg/ml blasticidin.
 Analyze your transformants for the presence of the insert by restriction digestion.
 Select a transformant with the correct restriction pattern and use sequencing to confirm that your gene is cloned in-frame with the C-terminal peptide.
 Transfect your construct into the cell line of choice using your own method of transfection.
  Test for expression of your recombinant gene by western blot analysis or functional assay. For antibodies to the myc epitope or the C-terminal polyhistidine tag, see the next page.
  To purify your recombinant protein, you may use metal-chelating resin such as ProBond. ProBond. resin is available separately.


载体序列

LOCUS       pcDNA6/myc-His C	5122 bp 	DNA	SYN
DEFINITION  pcDNA6/myc-His C
ACCESSION   
KEYWORDS    
SOURCE      
  ORGANISM  other sequences; artificial sequences; vectors.
FEATURES             Location/Qualifiers
     source          1..5122
                     /organism="pcDNA6/myc-His C"
                     /mol_type="other DNA"
     promoter        236..852
                     /label="CMV_immearly_promoter"
     misc_feature    315..602
                     /label="CAG_enhancer"
     misc_feature    769..789
                     /label="CMV_fwd_primer"
     promoter        863..881
                     /label="T7_promoter"
     misc_feature    complement(1078..1095)
                     /label="BGH_rev_primer"
     terminator      1081..1308
                     /label="bGH_PA_terminator"
     rep_origin      1371..1677
                     /label="f1_origin"
     misc_feature    complement(1791..1811)
                     /label="pBABE_3_primer"
     misc_feature    complement(1797..2012)
                     /label="SV40_enhancer"
     promoter        1809..2077
                     /label="SV40_promoter"
     rep_origin      1976..2053
                     /label="SV40_origin"
     misc_feature    2038..2057
                     /label="SV40pro_F_primer"
     promoter        2171..2238
                     /label="EM7_promoter"
     gene            2239..2637
                     /label="blast"
                     /gene="blast"
     terminator      2798..2917
                     /label="SV40_PA_terminator"
     misc_feature    2886..2905
                     /label="EBV_rev_primer"
     promoter        complement(2961..2979)
                     /label="M13_reverse_primer"
     misc_feature    complement(2978..3000)
                     /label="M13_pUC_rev_primer"
     promoter        complement(3014..3043)
                     /label="lac_promoter"
     gene            complement(4126..4986)
                     /label="Ampicillin"
                     /gene="Ampicillin"
     CDS             complement(4126..4986)
                     /label="ORF frame 2"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY
                     IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVE
                     YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL
                     DRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL
                     LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA
                     EIGASLIKHW*"
     promoter        complement(5028..5056)
                     /label="AmpR_promoter"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGY
                     IELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVE
                     YSPVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRL
                     DRWEPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPL
                     LRSALPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIA
                     EIGASLIKHW*"
ORIGIN
    1 GACGGATCGG GAGATCTCCC GATCCCCTAT GGTGCACTCT CAGTACAATC TGCTCTGATG
   61 CCGCATAGTT AAGCCAGTAT CTGCTCCCTG CTTGTGTGTT GGAGGTCGCT GAGTAGTGCG
  121 CGAGCAAAAT TTAAGCTACA ACAAGGCAAG GCTTGACCGA CAATTGCATG AAGAATCTGC
  181 TTAGGGTTAG GCGTTTTGCG CTGCTTCGCG ATGTACGGGC CAGATATACG CGTTGACATT
  241 GATTATTGAC TAGTTATTAA TAGTAATCAA TTACGGGGTC ATTAGTTCAT AGCCCATATA
  301 TGGAGTTCCG CGTTACATAA CTTACGGTAA ATGGCCCGCC TGGCTGACCG CCCAACGACC
  361 CCCGCCCATT GACGTCAATA ATGACGTATG TTCCCATAGT AACGCCAATA GGGACTTTCC
  421 ATTGACGTCA ATGGGTGGAG TATTTACGGT AAACTGCCCA CTTGGCAGTA CATCAAGTGT
  481 ATCATATGCC AAGTACGCCC CCTATTGACG TCAATGACGG TAAATGGCCC GCCTGGCATT
  541 ATGCCCAGTA CATGACCTTA TGGGACTTTC CTACTTGGCA GTACATCTAC GTATTAGTCA
  601 TCGCTATTAC CATGGTGATG CGGTTTTGGC AGTACATCAA TGGGCGTGGA TAGCGGTTTG
  661 ACTCACGGGG ATTTCCAAGT CTCCACCCCA TTGACGTCAA TGGGAGTTTG TTTTGGCACC
  721 AAAATCAACG GGACTTTCCA AAATGTCGTA ACAACTCCGC CCCATTGACG CAAATGGGCG
  781 GTAGGCGTGT ACGGTGGGAG GTCTATATAA GCAGAGCTCT CTGGCTAACT AGAGAACCCA
  841 CTGCTTACTG GCTTATCGAA ATTAATACGA CTCACTATAG GGAGACCCAA GCTGGCTAGT
  901 TAAGCTTGGT ACCGAGCTCG GATCCACTAG TCCAGTGTGG TGGAATTCTG CAGATATCCA
  961 GCACAGTGGC GGCCGCTCGA GGTCACCCAT TCGAACAAAA ACTCATCTCA GAAGAGGATC
 1021 TGAATATGCA TACCGGTCAT CATCACCATC ACCATTGAGT TTAAACCCGC TGATCAGCCT
 1081 CGACTGTGCC TTCTAGTTGC CAGCCATCTG TTGTTTGCCC CTCCCCCGTG CCTTCCTTGA
 1141 CCCTGGAAGG TGCCACTCCC ACTGTCCTTT CCTAATAAAA TGAGGAAATT GCATCGCATT
 1201 GTCTGAGTAG GTGTCATTCT ATTCTGGGGG GTGGGGTGGG GCAGGACAGC AAGGGGGAGG
 1261 ATTGGGAAGA CAATAGCAGG CATGCTGGGG ATGCGGTGGG CTCTATGGCT TCTGAGGCGG
 1321 AAAGAACCAG CTGGGGCTCT AGGGGGTATC CCCACGCGCC CTGTAGCGGC GCATTAAGCG
 1381 CGGCGGGTGT GGTGGTTACG CGCAGCGTGA CCGCTACACT TGCCAGCGCC CTAGCGCCCG
 1441 CTCCTTTCGC TTTCTTCCCT TCCTTTCTCG CCACGTTCGC CGGCTTTCCC CGTCAAGCTC
 1501 TAAATCGGGG GCTCCCTTTA GGGTTCCGAT TTAGTGCTTT ACGGCACCTC GACCCCAAAA
 1561 AACTTGATTA GGGTGATGGT TCACGTAGTG GGCCATCGCC CTGATAGACG GTTTTTCGCC
 1621 CTTTGACGTT GGAGTCCACG TTCTTTAATA GTGGACTCTT GTTCCAAACT GGAACAACAC
 1681 TCAACCCTAT CTCGGTCTAT TCTTTTGATT TATAAGGGAT TTTGCCGATT TCGGCCTATT
 1741 GGTTAAAAAA TGAGCTGATT TAACAAAAAT TTAACGCGAA TTAATTCTGT GGAATGTGTG
 1801 TCAGTTAGGG TGTGGAAAGT CCCCAGGCTC CCCAGCAGGC AGAAGTATGC AAAGCATGCA
 1861 TCTCAATTAG TCAGCAACCA GGTGTGGAAA GTCCCCAGGC TCCCCAGCAG GCAGAAGTAT
 1921 GCAAAGCATG CATCTCAATT AGTCAGCAAC CATAGTCCCG CCCCTAACTC CGCCCATCCC
 1981 GCCCCTAACT CCGCCCAGTT CCGCCCATTC TCCGCCCCAT GGCTGACTAA TTTTTTTTAT
 2041 TTATGCAGAG GCCGAGGCCG CCTCTGCCTC TGAGCTATTC CAGAAGTAGT GAGGAGGCTT
 2101 TTTTGGAGGC CTAGGCTTTT GCAAAAAGCT CCCGGGAGCT TGTATATCCA TTTTCGGATC
 2161 TGATCAGCAC GTGTTGACAA TTAATCATCG GCATAGTATA TCGGCATAGT ATAATACGAC
 2221 AAGGTGAGGA ACTAAACCAT GGCCAAGCCT TTGTCTCAAG AAGAATCCAC CCTCATTGAA
 2281 AGAGCAACGG CTACAATCAA CAGCATCCCC ATCTCTGAAG ACTACAGCGT CGCCAGCGCA
 2341 GCTCTCTCTA GCGACGGCCG CATCTTCACT GGTGTCAATG TATATCATTT TACTGGGGGA
 2401 CCTTGTGCAG AACTCGTGGT GCTGGGCACT GCTGCTGCTG CGGCAGCTGG CAACCTGACT
 2461 TGTATCGTCG CGATCGGAAA TGAGAACAGG GGCATCTTGA GCCCCTGCGG ACGGTGCCGA
 2521 CAGGTGCTTC TCGATCTGCA TCCTGGGATC AAAGCCATAG TGAAGGACAG TGATGGACAG
 2581 CCGACGGCAG TTGGGATTCG TGAATTGCTG CCCTCTGGTT ATGTGTGGGA GGGCTAAGCA
 2641 CTTCGTGGCC GAGGAGCAGG ACTGACACGT GCTACGAGAT TTCGATTCCA CCGCCGCCTT
 2701 CTATGAAAGG TTGGGCTTCG GAATCGTTTT CCGGGACGCC GGCTGGATGA TCCTCCAGCG
 2761 CGGGGATCTC ATGCTGGAGT TCTTCGCCCA CCCCAACTTG TTTATTGCAG CTTATAATGG
 2821 TTACAAATAA AGCAATAGCA TCACAAATTT CACAAATAAA GCATTTTTTT CACTGCATTC
 2881 TAGTTGTGGT TTGTCCAAAC TCATCAATGT ATCTTATCAT GTCTGTATAC CGTCGACCTC
 2941 TAGCTAGAGC TTGGCGTAAT CATGGTCATA GCTGTTTCCT GTGTGAAATT GTTATCCGCT
 3001 CACAATTCCA CACAACATAC GAGCCGGAAG CATAAAGTGT AAAGCCTGGG GTGCCTAATG
 3061 AGTGAGCTAA CTCACATTAA TTGCGTTGCG CTCACTGCCC GCTTTCCAGT CGGGAAACCT
 3121 GTCGTGCCAG CTGCATTAAT GAATCGGCCA ACGCGCGGGG AGAGGCGGTT TGCGTATTGG
 3181 GCGCTCTTCC GCTTCCTCGC TCACTGACTC GCTGCGCTCG GTCGTTCGGC TGCGGCGAGC
 3241 GGTATCAGCT CACTCAAAGG CGGTAATACG GTTATCCACA GAATCAGGGG ATAACGCAGG
 3301 AAAGAACATG TGAGCAAAAG GCCAGCAAAA GGCCAGGAAC CGTAAAAAGG CCGCGTTGCT
 3361 GGCGTTTTTC CATAGGCTCC GCCCCCCTGA CGAGCATCAC AAAAATCGAC GCTCAAGTCA
 3421 GAGGTGGCGA AACCCGACAG GACTATAAAG ATACCAGGCG TTTCCCCCTG GAAGCTCCCT
 3481 CGTGCGCTCT CCTGTTCCGA CCCTGCCGCT TACCGGATAC CTGTCCGCCT TTCTCCCTTC
 3541 GGGAAGCGTG GCGCTTTCTC ATAGCTCACG CTGTAGGTAT CTCAGTTCGG TGTAGGTCGT
 3601 TCGCTCCAAG CTGGGCTGTG TGCACGAACC CCCCGTTCAG CCCGACCGCT GCGCCTTATC
 3661 CGGTAACTAT CGTCTTGAGT CCAACCCGGT AAGACACGAC TTATCGCCAC TGGCAGCAGC
 3721 CACTGGTAAC AGGATTAGCA GAGCGAGGTA TGTAGGCGGT GCTACAGAGT TCTTGAAGTG
 3781 GTGGCCTAAC TACGGCTACA CTAGAAGAAC AGTATTTGGT ATCTGCGCTC TGCTGAAGCC
 3841 AGTTACCTTC GGAAAAAGAG TTGGTAGCTC TTGATCCGGC AAACAAACCA CCGCTGGTAG
 3901 CGGTGGTTTT TTTGTTTGCA AGCAGCAGAT TACGCGCAGA AAAAAAGGAT CTCAAGAAGA
 3961 TCCTTTGATC TTTTCTACGG GGTCTGACGC TCAGTGGAAC GAAAACTCAC GTTAAGGGAT
 4021 TTTGGTCATG AGATTATCAA AAAGGATCTT CACCTAGATC CTTTTAAATT AAAAATGAAG
 4081 TTTTAAATCA ATCTAAAGTA TATATGAGTA AACTTGGTCT GACAGTTACC AATGCTTAAT
 4141 CAGTGAGGCA CCTATCTCAG CGATCTGTCT ATTTCGTTCA TCCATAGTTG CCTGACTCCC
 4201 CGTCGTGTAG ATAACTACGA TACGGGAGGG CTTACCATCT GGCCCCAGTG CTGCAATGAT
 4261 ACCGCGAGAC CCACGCTCAC CGGCTCCAGA TTTATCAGCA ATAAACCAGC CAGCCGGAAG
 4321 GGCCGAGCGC AGAAGTGGTC CTGCAACTTT ATCCGCCTCC ATCCAGTCTA TTAATTGTTG
 4381 CCGGGAAGCT AGAGTAAGTA GTTCGCCAGT TAATAGTTTG CGCAACGTTG TTGCCATTGC
 4441 TACAGGCATC GTGGTGTCAC GCTCGTCGTT TGGTATGGCT TCATTCAGCT CCGGTTCCCA
 4501 ACGATCAAGG CGAGTTACAT GATCCCCCAT GTTGTGCAAA AAAGCGGTTA GCTCCTTCGG
 4561 TCCTCCGATC GTTGTCAGAA GTAAGTTGGC CGCAGTGTTA TCACTCATGG TTATGGCAGC
 4621 ACTGCATAAT TCTCTTACTG TCATGCCATC CGTAAGATGC TTTTCTGTGA CTGGTGAGTA
 4681 CTCAACCAAG TCATTCTGAG AATAGTGTAT GCGGCGACCG AGTTGCTCTT GCCCGGCGTC
 4741 AATACGGGAT AATACCGCGC CACATAGCAG AACTTTAAAA GTGCTCATCA TTGGAAAACG
 4801 TTCTTCGGGG CGAAAACTCT CAAGGATCTT ACCGCTGTTG AGATCCAGTT CGATGTAACC
 4861 CACTCGTGCA CCCAACTGAT CTTCAGCATC TTTTACTTTC ACCAGCGTTT CTGGGTGAGC
 4921 AAAAACAGGA AGGCAAAATG CCGCAAAAAA GGGAATAAGG GCGACACGGA AATGTTGAAT
 4981 ACTCATACTC TTCCTTTTTC AATATTATTG AAGCATTTAT CAGGGTTATT GTCTCATGAG
 5041 CGGATACATA TTTGAATGTA TTTAGAAAAA TAAACAAATA GGGGTTCCGC GCACATTTCC
 5101 CCGAAAAGTG CCACCTGACG TC
//

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