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          pDsRED-Monomer-N1

          價格:1200元

          聯系方式:I47-825O-882O

          買家導航

          載體基本信息

          出品公司: Clontech
          載體名稱: pDsRed-Monomer-N1
          質粒類型: 哺乳動物細胞表達載體;熒光報告載體
          高拷貝/低拷貝: 高拷貝
          啟動子: CMV
          克隆方法: 多克隆位點,限制性內切酶
          載體大小: 4691bp
          5' 測序引物及序列: 5'd[GTACTGGAACTGGGGGGACAG]
          3' 測序引物及序列: --
          載體標簽: 紅色熒光蛋白DsRed-Monomer (C-term)
          載體抗性: Kanamycin
          篩選標記: Neomycin
          克隆菌株: DH5α, HB101
          宿主細胞(系): 常規細胞系,293、CV-1、CHO等
          備注:
          pDsRed-Monomer-N1載體是熒光報告載體,表達C端DsRed-Monomer融合蛋白;
          DsRed-Monomer是DsRed的單體突變體,與DsRed相比,編碼序列經過了人工優化,適用于在哺乳動物細胞中
          的高水平表達。
          產品目錄號: 632465
          穩定性: 穩表達 或 瞬表達
          組成型: 組成型
          病毒/非病毒: 非病毒

          載體質粒圖譜和多克隆位點信息

          pDsRED-Monomer-N1載體圖譜
          pDsRED-Monomer-N1多克隆位點

          pDsRED-Monomer-N1載體特征

          載體簡介

          pDsRed-Express-N1載體描述
          
          pDsRed-Express-N1 is a mammalian expression vector that encodes DsRed-Express, a variant of Discosoma sp. red fluorescent protein (DsRed; 1). DsRed-Express contains nine amino acid substitutions (listed on page 2), which improve the solubility of the protein and reduce the time from transfection to detection of red fluorescence (2). In addition, these substitutions reduce the level of residual green emission (2). When DsRed-Express is expressed in mammalian cell cultures, redemitting cells can be detected by either fluorescence microscopy or flow cytometry 8–12 hours after transfection (DsRed-Express excitation and emission maxima = 557 nm and 579 nm, respectively). Although DsRed-Express most likely forms the same tetrameric structure as wild-type DsRed, DsRed- Express displays a reduced tendency to aggregate (2). The DsRed-Express coding sequence is human codon-optimized for high expression in mammalian cells (3).
          The multiple cloning site (MCS) in pDsRed-Express-N1 is positioned between the immediate early promoter of CMV (PCMV IE) and the DsRed-Express coding sequence. Genes cloned into the MCS are expressed as fusions to the N-terminus of DsRed-Express if they are in the same reading frame as DsRed-Express and there are no intervening stop codons. Sequences upstream of DsRed-Express have been converted to a Kozak consensus translation initiation site to increase translation efficiency in eukaryotic cells (4). SV40 polyadenylation signals downstream of the DsRed-Express gene direct proper processing of the 3' end of the DsRed-Express mRNA. The vector backbone contains an SV40 origin for replication in mammalian cells expressing the SV40 T antigen, a pUC origin of replication for propagation in E. coli, and an f1 origin for single-stranded DNA production. A neomycin-resistance cassette (Neor) allows stably transfected eukaryotic cells to be selected using G418. This cassette consists of the SV40 early promoter, the neomycin/kanamycin resistance gene of Tn5, and polyadenylation signals from the Herpes simplex virus thymidine kinase (HSV TK) gene.A bacterial promoter upstream of the cassette confers kanamycin resistance to E. coli.
          
          
          pDsRed-Express-N1 can be used to construct fusions to the N-terminus of DsRed-Express. If a fusion construct retains the fluorescent properties of the native DsRed-Express protein, its expression can be monitored by flow cytometry and its localization in vivo can be determined by fluorescence microscopy. The target gene should be cloned into pDsRed-Express-N1 so that it is in frame with the DsRed-Express coding sequence, with no intervening in-frame stop codons. The inserted gene should include an initiating ATG codon. Recombinant pDsRed-Express-N1 can be transfected into mammalian cells using any standard transfection method. If required, stable transfectants can be selected using G418 (5). pDsRed-Express-N1 can also be used as a cotransfection marker; the unmodified vector will express DsRed-Express.
          
          
          pDsRed-Express-N1載體含有以下元件:
          
           Human cytomegalovirus (CMV) immediate early promoter: 1–589
          Enhancer region: 59–465; TATA box: 554–560
          Transcription start point: 583
          C→G mutation to remove Sac I site: 569
           MCS: 591–671
           Discosoma sp. Red Fluorescent Protein (DsRed-Express) gene
          Kozak consensus translation initiation site: 672–682
          Start codon (ATG): 679–681; Stop codon: 1354–1356
          CGC→GCC (Arg-2 to Ala) mutation: 682–684
          AAG→GAG (Lys-5 to Glu) mutation: 691–693
          AAC→GAC (Asn-6 to Asp) mutation: 694–696
          ACC→TCC (Thr-21 to Ser) mutation: 739–741
          CAC→ACC (His-41 to Thr) mutation: 799–801
          AAC→CAG (Asn-42 to Gln) mutation: 802–804
          GTG→GCC (Val-44 to Ala) mutation: 808–810
          TGC→TCC (Cys-117 to Ser) mutation: 1027–1029
          ACC→GCC (Thr-217 to Ala) mutation: 1327–1329
           SV40 early mRNA polyadenylation signal
          Polyadenylation signals: 1508–1513 & 1537–1542; mRNA 3' ends: 1546 & 1558
           f1 single-strand DNA origin: 1605–2060 (Packages the noncoding strand of DsRed-Express.)
           Bacterial promoter for expression of Kanr gene:
          –35 region: 2122–2127; –10 region: 2145–2150
          Transcription start point: 2157
           SV40 origin of replication: 2401–2536
           SV40 early promoter
          Enhancer (72-bp tandem repeats): 2234–2305 & 2306–2377
          21-bp repeats: 2381–2401, 2402–2422 & 2424–2444
          Early promoter element: 2457–2463
          Major transcription start points: 2453, 2491, 2497 & 2502
           Kanamycin/neomycin resistance gene
          Neomycin phosphotransferase coding sequences: Start codon (ATG): 2585–2587; Stop codon: 3377–3379
          G→A mutation to remove Pst I site: 2767
          C→A (Arg to Ser) mutation to remove BssH II site: 3113
           Herpes simplex virus (HSV) thymidine kinase (TK) polyadenylation signal
          Polyadenylation signals: 3615–3620 & 3628–3633
           pUC plasmid replication origin: 3964–4607
          Sequencing primer locations
           DsRed1-N Sequencing Primer (Cat. No. 632387; 5'-GTACTGGAACTGGGGGGACAG-3'): 879–859
          
          Propagation in E. coli
           Suitable host strains: DH5α, HB101 and other general purpose strains. Single-stranded DNA production requires
          a host containing an F plasmid, such as the JM109 or XL1-Blue strains..
           Selectable marker: plasmid confers resistance to kanamycin (50 μg/ml) to E. coli hosts.
           E. coli replication origin: pUC
           Copy number: ≈500
           Plasmid incompatibility group: pMB1/ColE1
          
          Excitation and emission maxima of DsRed-Express
           Excitation maximum = 557 nm
           Emission maximum = 579 nm
          

          載體序列

          LOCUS       pDsRed-Monomer-N1	4691 bp 	DNA	SYN
          DEFINITION  pDsRed-Monomer-N1
          ACCESSION   
          KEYWORDS    
          SOURCE      
            ORGANISM  other sequences; artificial sequences; vectors.
          FEATURES             Location/Qualifiers
               source          1..4691
                               /organism="pDsRed-Monomer-N1"
                               /mol_type="other DNA"
               promoter        10..562
                               /label="CMV_immearly_promoter"
               misc_feature    65..352
                               /label="CAG_enhancer"
               misc_feature    519..539
                               /label="CMV_fwd_primer"
               promoter        520..589
                               /label="CMV_promoter"
               CDS             679..1356
                               /label="ORF frame 1"
          
               gene            682..1353
                               /label="DsRed_Monomer"
                               /gene="DsRed_Monomer"
          
               misc_feature    complement(859..879)
                               /label="dsRed1_N_primer"
          
               misc_feature    1570..1589
                               /label="EBV_rev_primer"
          
               rep_origin      complement(1739..2045)
                               /label="f1_origin"
          
               promoter        2124..2152
                               /label="AmpR_promoter"
          
               misc_feature    complement(2218..2238)
                               /label="pBABE_3_primer"
          
               misc_feature    complement(2224..2439)
                               /label="SV40_enhancer"
          
               promoter        2236..2504
                               /label="SV40_promoter"
          
               rep_origin      2403..2480
                               /label="SV40_origin"
          
               misc_feature    2465..2484
                               /label="SV40pro_F_primer"
          
               CDS             2587..3381
                               /label="ORF frame 1"
          
               gene            2590..3378
                               /label="NeoR/KanR"
                               /gene="NeoR/KanR"
          
               CDS             complement(2896..3432)
                               /label="ORF frame 3"
          
               terminator      3556..3825
                               /label="TK_PA_terminator"
          
               rep_origin      3973..4592
                               /label="pBR322_origin"
          
          ORIGIN
              1 TAGTTATTAA TAGTAATCAA TTACGGGGTC ATTAGTTCAT AGCCCATATA TGGAGTTCCG
             61 CGTTACATAA CTTACGGTAA ATGGCCCGCC TGGCTGACCG CCCAACGACC CCCGCCCATT
            121 GACGTCAATA ATGACGTATG TTCCCATAGT AACGCCAATA GGGACTTTCC ATTGACGTCA
            181 ATGGGTGGAG TATTTACGGT AAACTGCCCA CTTGGCAGTA CATCAAGTGT ATCATATGCC
            241 AAGTACGCCC CCTATTGACG TCAATGACGG TAAATGGCCC GCCTGGCATT ATGCCCAGTA
            301 CATGACCTTA TGGGACTTTC CTACTTGGCA GTACATCTAC GTATTAGTCA TCGCTATTAC
            361 CATGGTGATG CGGTTTTGGC AGTACATCAA TGGGCGTGGA TAGCGGTTTG ACTCACGGGG
            421 ATTTCCAAGT CTCCACCCCA TTGACGTCAA TGGGAGTTTG TTTTGGCACC AAAATCAACG
            481 GGACTTTCCA AAATGTCGTA ACAACTCCGC CCCATTGACG CAAATGGGCG GTAGGCGTGT
            541 ACGGTGGGAG GTCTATATAA GCAGAGCTGG TTTAGTGAAC CGTCAGATCC GCTAGCGCTA
            601 CCGGACTCAG ATCTCGAGCT CAAGCTTCGA ATTCTGCAGT CGACGGTACC GCGGGCCCGG
            661 GATCCACCGG TCGCCACCAT GGACAACACC GAGGACGTCA TCAAGGAGTT CATGCAGTTC
            721 AAGGTGCGCA TGGAGGGCTC CGTGAACGGC CACTACTTCG AGATCGAGGG CGAGGGCGAG
            781 GGCAAGCCCT ACGAGGGCAC CCAGACCGCC AAGCTGCAGG TGACCAAGGG CGGCCCCCTG
            841 CCCTTCGCCT GGGACATCCT GTCCCCCCAG TTCCAGTACG GCTCCAAGGC CTACGTGAAG
            901 CACCCCGCCG ACATCCCCGA CTACATGAAG CTGTCCTTCC CCGAGGGCTT CACCTGGGAG
            961 CGCTCCATGA ACTTCGAGGA CGGCGGCGTG GTGGAGGTGC AGCAGGACTC CTCCCTGCAG
           1021 GACGGCACCT TCATCTACAA GGTGAAGTTC AAGGGCGTGA ACTTCCCCGC CGACGGCCCC
           1081 GTAATGCAGA AGAAGACTGC CGGCTGGGAG CCCTCCACCG AGAAGCTGTA CCCCCAGGAC
           1141 GGCGTGCTGA AGGGCGAGAT CTCCCACGCC CTGAAGCTGA AGGACGGCGG CCACTACACC
           1201 TGCGACTTCA AGACCGTGTA CAAGGCCAAG AAGCCCGTGC AGCTGCCCGG CAACCACTAC
           1261 GTGGACTCCA AGCTGGACAT CACCAACCAC AACGAGGACT ACACCGTGGT GGAGCAGTAC
           1321 GAGCACGCCG AGGCCCGCCA CTCCGGCTCC CAGTAGAGCG GCCGCGACTC TAGATCATAA
           1381 TCAGCCATAC CACATTTGTA GAGGTTTTAC TTGCTTTAAA AAACCTCCCA CACCTCCCCC
           1441 TGAACCTGAA ACATAAAATG AATGCAATTG TTGTTGTTAA CTTGTTTATT GCAGCTTATA
           1501 ATGGTTACAA ATAAAGCAAT AGCATCACAA ATTTCACAAA TAAAGCATTT TTTTCACTGC
           1561 ATTCTAGTTG TGGTTTGTCC AAACTCATCA ATGTATCTTA AGGCGTAAAT TGTAAGCGTT
           1621 AATATTTTGT TAAAATTCGC GTTAAATTTT TGTTAAATCA GCTCATTTTT TAACCAATAG
           1681 GCCGAAATCG GCAAAATCCC TTATAAATCA AAAGAATAGA CCGAGATAGG GTTGAGTGTT
           1741 GTTCCAGTTT GGAACAAGAG TCCACTATTA AAGAACGTGG ACTCCAACGT CAAAGGGCGA
           1801 AAAACCGTCT ATCAGGGCGA TGGCCCACTA CGTGAACCAT CACCCTAATC AAGTTTTTTG
           1861 GGGTCGAGGT GCCGTAAAGC ACTAAATCGG AACCCTAAAG GGAGCCCCCG ATTTAGAGCT
           1921 TGACGGGGAA AGCCGGCGAA CGTGGCGAGA AAGGAAGGGA AGAAAGCGAA AGGAGCGGGC
           1981 GCTAGGGCGC TGGCAAGTGT AGCGGTCACG CTGCGCGTAA CCACCACACC CGCCGCGCTT
           2041 AATGCGCCGC TACAGGGCGC GTCAGGTGGC ACTTTTCGGG GAAATGTGCG CGGAACCCCT
           2101 ATTTGTTTAT TTTTCTAAAT ACATTCAAAT ATGTATCCGC TCATGAGACA ATAACCCTGA
           2161 TAAATGCTTC AATAATATTG AAAAAGGAAG AGTCCTGAGG CGGAAAGAAC CAGCTGTGGA
           2221 ATGTGTGTCA GTTAGGGTGT GGAAAGTCCC CAGGCTCCCC AGCAGGCAGA AGTATGCAAA
           2281 GCATGCATCT CAATTAGTCA GCAACCAGGT GTGGAAAGTC CCCAGGCTCC CCAGCAGGCA
           2341 GAAGTATGCA AAGCATGCAT CTCAATTAGT CAGCAACCAT AGTCCCGCCC CTAACTCCGC
           2401 CCATCCCGCC CCTAACTCCG CCCAGTTCCG CCCATTCTCC GCCCCATGGC TGACTAATTT
           2461 TTTTTATTTA TGCAGAGGCC GAGGCCGCCT CGGCCTCTGA GCTATTCCAG AAGTAGTGAG
           2521 GAGGCTTTTT TGGAGGCCTA GGCTTTTGCA AAGATCGATC AAGAGACAGG ATGAGGATCG
           2581 TTTCGCATGA TTGAACAAGA TGGATTGCAC GCAGGTTCTC CGGCCGCTTG GGTGGAGAGG
           2641 CTATTCGGCT ATGACTGGGC ACAACAGACA ATCGGCTGCT CTGATGCCGC CGTGTTCCGG
           2701 CTGTCAGCGC AGGGGCGCCC GGTTCTTTTT GTCAAGACCG ACCTGTCCGG TGCCCTGAAT
           2761 GAACTGCAAG ACGAGGCAGC GCGGCTATCG TGGCTGGCCA CGACGGGCGT TCCTTGCGCA
           2821 GCTGTGCTCG ACGTTGTCAC TGAAGCGGGA AGGGACTGGC TGCTATTGGG CGAAGTGCCG
           2881 GGGCAGGATC TCCTGTCATC TCACCTTGCT CCTGCCGAGA AAGTATCCAT CATGGCTGAT
           2941 GCAATGCGGC GGCTGCATAC GCTTGATCCG GCTACCTGCC CATTCGACCA CCAAGCGAAA
           3001 CATCGCATCG AGCGAGCACG TACTCGGATG GAAGCCGGTC TTGTCGATCA GGATGATCTG
           3061 GACGAAGAGC ATCAGGGGCT CGCGCCAGCC GAACTGTTCG CCAGGCTCAA GGCGAGCATG
           3121 CCCGACGGCG AGGATCTCGT CGTGACCCAT GGCGATGCCT GCTTGCCGAA TATCATGGTG
           3181 GAAAATGGCC GCTTTTCTGG ATTCATCGAC TGTGGCCGGC TGGGTGTGGC GGACCGCTAT
           3241 CAGGACATAG CGTTGGCTAC CCGTGATATT GCTGAAGAGC TTGGCGGCGA ATGGGCTGAC
           3301 CGCTTCCTCG TGCTTTACGG TATCGCCGCT CCCGATTCGC AGCGCATCGC CTTCTATCGC
           3361 CTTCTTGACG AGTTCTTCTG AGCGGGACTC TGGGGTTCGA AATGACCGAC CAAGCGACGC
           3421 CCAACCTGCC ATCACGAGAT TTCGATTCCA CCGCCGCCTT CTATGAAAGG TTGGGCTTCG
           3481 GAATCGTTTT CCGGGACGCC GGCTGGATGA TCCTCCAGCG CGGGGATCTC ATGCTGGAGT
           3541 TCTTCGCCCA CCCTAGGGGG AGGCTAACTG AAACACGGAA GGAGACAATA CCGGAAGGAA
           3601 CCCGCGCTAT GACGGCAATA AAAAGACAGA ATAAAACGCA CGGTGTTGGG TCGTTTGTTC
           3661 ATAAACGCGG GGTTCGGTCC CAGGGCTGGC ACTCTGTCGA TACCCCACCG AGACCCCATT
           3721 GGGGCCAATA CGCCCGCGTT TCTTCCTTTT CCCCACCCCA CCCCCCAAGT TCGGGTGAAG
           3781 GCCCAGGGCT CGCAGCCAAC GTCGGGGCGG CAGGCCCTGC CATAGCCTCA GGTTACTCAT
           3841 ATATACTTTA GATTGATTTA AAACTTCATT TTTAATTTAA AAGGATCTAG GTGAAGATCC
           3901 TTTTTGATAA TCTCATGACC AAAATCCCTT AACGTGAGTT TTCGTTCCAC TGAGCGTCAG
           3961 ACCCCGTAGA AAAGATCAAA GGATCTTCTT GAGATCCTTT TTTTCTGCGC GTAATCTGCT
           4021 GCTTGCAAAC AAAAAAACCA CCGCTACCAG CGGTGGTTTG TTTGCCGGAT CAAGAGCTAC
           4081 CAACTCTTTT TCCGAAGGTA ACTGGCTTCA GCAGAGCGCA GATACCAAAT ACTGTCCTTC
           4141 TAGTGTAGCC GTAGTTAGGC CACCACTTCA AGAACTCTGT AGCACCGCCT ACATACCTCG
           4201 CTCTGCTAAT CCTGTTACCA GTGGCTGCTG CCAGTGGCGA TAAGTCGTGT CTTACCGGGT
           4261 TGGACTCAAG ACGATAGTTA CCGGATAAGG CGCAGCGGTC GGGCTGAACG GGGGGTTCGT
           4321 GCACACAGCC CAGCTTGGAG CGAACGACCT ACACCGAACT GAGATACCTA CAGCGTGAGC
           4381 TATGAGAAAG CGCCACGCTT CCCGAAGGGA GAAAGGCGGA CAGGTATCCG GTAAGCGGCA
           4441 GGGTCGGAAC AGGAGAGCGC ACGAGGGAGC TTCCAGGGGG AAACGCCTGG TATCTTTATA
           4501 GTCCTGTCGG GTTTCGCCAC CTCTGACTTG AGCGTCGATT TTTGTGATGC TCGTCAGGGG
           4561 GGCGGAGCCT ATGGAAAAAC GCCAGCAACG CGGCCTTTTT ACGGTTCCTG GCCTTTTGCT
           4621 GGCCTTTTGC TCACATGTTC TTTCCTGCGT TATCCCCTGA TTCTGTGGAT AACCGTATTA
           4681 CCGCCATGCA T
          //
          

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