pAsRed2-N1载体

pAsRed2-N1载体

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  • ¥2300
  • ZYbscience
  • 中国/美国
  • ZY
  • 2025年07月14日
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    上海泽叶生物科技有限公司
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    • 详细信息
    • 技术资料
    • 保存条件：

      -20℃低温保存

    • 保质期：

      三年

    • 英文名：

      pAsRed2-N1

    • 库存：

      20

    • 供应商：

      泽叶生物

    载体基本信息

    出品公司: ZYbscience
    载体名称: pAsRed2-N1
    质粒类型: 哺乳动物细胞表达载体；荧光报告载体
    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 IE
    载体大小: 4712 bp
    5' 测序引物及序列: --
    3' 测序引物及序列: --
    载体标签: AsRed2 (C-端）
    载体抗性: 卡那霉素
    筛选标记: 新霉素（Neomycin）
    克隆菌株: DH5α, HB101
    宿主细胞（系）: 常规细胞系（293、CV-1、CHO等）
    备注: 哺乳动物载体pAsRed2-N1组成型表达C端pAsRed2融合蛋白。
    产品目录号: ZY632449
    稳定性: 稳表达 或 瞬表达
    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

    pAsRed2-N1载体图谱



    pAsRed2-N1 多克隆位点

    pAsRed2-N1 载体特征1
    pAsRed2-N1 载体特征2

    载体简介

    
pAsRed2-N1 encodes AsRed2, a variant of Anemonia sulcata red fluorescent protein (1, 2). AsRed2 has been engineered for brighter fluorescence (Clontech Laboratories, Inc., unpublished data). The AsRed2 coding sequence also contains a series of silent base-pair changes, which correspond to human codon-usage preferences, for optimal expression in mammalian cells (3). Additionally, an upstream sequence—located just 5' to the AsRed2 start codon—has been converted to a Kozak consensus translation initiation site (4) to further increase the translation efficiency in eukaryotic cells.

The multiple cloning site (MCS) in pAsRed2-N1 is positioned between the immediate-early promoter of cytomegalovirus (PCMV IE) and the AsRed2 coding sequence. Thus, genes cloned into the MCS will be expressed as fusions to the N-terminus of AsRed2 if they are in the same reading frame as AsRed2 and there are no intervening stop codons. The SV40 polyadenylation signals (SV40 poly A) downstream of the AsRed2 gene direct proper processing of the 3' end of AsRed2 mRNA. The vector backbone contains an SV40 origin (SV40 ori) for replication in mammalian cells that express the SV40 T antigen, a pUC origin of replication (pUC ori) for propagation in E. coli, and an f1 origin (f1 ori) for single-stranded DNA production. In addition, a neomycin-resistance cassette—consisting of the SV40 early promoter (PSV40e), the neomycin/kanamycin resistance gene of Tn5 (Neor/Kanr), and polyadenylation signals from the Herpes simplex virus thymidine kinase(HSV TK poly A) gene—allows stably transfected eukaryotic cells to be selected using G418 (5). A bacterial promoter (P) upstream of this cassette drives expression of the Neor/Kanr gene in E. coli hosts, which can be selected with kanamycin.

Fusions to the N terminus of AsRed2 retain the fluorescent properties of the native protein allowing the localization of the fusion protein in vivo (AsRed2 excitation maximum = 576 nm; AsRed2 emission maximum = 592 nm). The target gene should be cloned into pAsRed2-N1 so that it is in frame with the AsRed2 coding sequence, with no intervening, in-frame stop codons. The inserted gene should include the initiating ATG codon. The recombinant pAsRed2-N1 vector can be transfected into mammalian cells using any standard transfection method. If required, stable transformants can be selected using G418 (available from Clontech; Cat. Nos. 631307 & 631308). We recommend selecting mammalian cell cultures in 500–1,300 μg/ml G418, depending on the cell line. Be sure to establish a kill curve for each cell line and each lot of G418 to determine the optimal selection concentration. Unmodified (i.e., non-recombinant) pAsRed2-N1 can also be used simply to express AsRed2 in cells of interest (e.g., as a transfection marker).

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 JM101 or XL1-Blue.
 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/Col E1

    载体序列

    LOCUS       pAsRed2-N1	4712 bp 	DNA	SYN
DEFINITION  pAsRed2-N1
ACCESSION   
KEYWORDS    
SOURCE      
  ORGANISM  other sequences; artificial sequences; vectors.
FEATURES             Location/Qualifiers
     source          1..4712
                     /organism="pAsRed2-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..1377
                     /label="ORF frame 1"
                     /translation="MASLLKKTMPFRTTIEGTVNGHYFKCTGKGEGNPLEGTQEMKIE
                     VIEGGPLPFAFHILSTSCMYGSKAFIKYVSGIPDYFKQSLPEGFTWERTTTYEDGGFL
                     TAHQDTSLDGDCLVYKVKILGNNFPADGPVMQNKAGRWEPSTEIVYEVDGVLRGQSLM
                     ALECPGGRHLTCHLHTTYRSKKPASALKMPGFHFEDHRIEILEEVEKGKCYKQYEAAV
                     GRYCDAAPSKLGHN*"
     gene            682..1365
                     /label="AsRed2 (variant)"
                     /gene="AsRed2 (variant)"
                     /translation="MASLLKKTMPFRTTIEGTVNGHYFKCTGKGEGNPLEGTQEMKIE
                     VIEGGPLPFAFHILSTSCMYGSKAFIKYVSGIPDYFKQSLPEGFTWERTTTYEDGGFL
                     TAHQDTSLDGDCLVYKVKILGNNFPADGPVMQNKAGRWEPSTEIVYEVDGVLRGQSLM
                     ALECPGGRHLTCHLHTTYRSKKPASALKMPGFHFEDHRIEILEEVEKGKCYKQYEAAV
                     GRYCDAAPSKLGHN*"
     misc_feature    1591..1610
                     /label="EBV_rev_primer"
                     /translation="MASLLKKTMPFRTTIEGTVNGHYFKCTGKGEGNPLEGTQEMKIE
                     VIEGGPLPFAFHILSTSCMYGSKAFIKYVSGIPDYFKQSLPEGFTWERTTTYEDGGFL
                     TAHQDTSLDGDCLVYKVKILGNNFPADGPVMQNKAGRWEPSTEIVYEVDGVLRGQSLM
                     ALECPGGRHLTCHLHTTYRSKKPASALKMPGFHFEDHRIEILEEVEKGKCYKQYEAAV
                     GRYCDAAPSKLGHN*"
     rep_origin      complement(1760..2066)
                     /label="f1_origin"
                     /translation="MASLLKKTMPFRTTIEGTVNGHYFKCTGKGEGNPLEGTQEMKIE
                     VIEGGPLPFAFHILSTSCMYGSKAFIKYVSGIPDYFKQSLPEGFTWERTTTYEDGGFL
                     TAHQDTSLDGDCLVYKVKILGNNFPADGPVMQNKAGRWEPSTEIVYEVDGVLRGQSLM
                     ALECPGGRHLTCHLHTTYRSKKPASALKMPGFHFEDHRIEILEEVEKGKCYKQYEAAV
                     GRYCDAAPSKLGHN*"
     promoter        2145..2173
                     /label="AmpR_promoter"
                     /translation="MASLLKKTMPFRTTIEGTVNGHYFKCTGKGEGNPLEGTQEMKIE
                     VIEGGPLPFAFHILSTSCMYGSKAFIKYVSGIPDYFKQSLPEGFTWERTTTYEDGGFL
                     TAHQDTSLDGDCLVYKVKILGNNFPADGPVMQNKAGRWEPSTEIVYEVDGVLRGQSLM
                     ALECPGGRHLTCHLHTTYRSKKPASALKMPGFHFEDHRIEILEEVEKGKCYKQYEAAV
                     GRYCDAAPSKLGHN*"
     misc_feature    complement(2239..2259)
                     /label="pBABE_3_primer"
                     /translation="MASLLKKTMPFRTTIEGTVNGHYFKCTGKGEGNPLEGTQEMKIE
                     VIEGGPLPFAFHILSTSCMYGSKAFIKYVSGIPDYFKQSLPEGFTWERTTTYEDGGFL
                     TAHQDTSLDGDCLVYKVKILGNNFPADGPVMQNKAGRWEPSTEIVYEVDGVLRGQSLM
                     ALECPGGRHLTCHLHTTYRSKKPASALKMPGFHFEDHRIEILEEVEKGKCYKQYEAAV
                     GRYCDAAPSKLGHN*"
     misc_feature    complement(2245..2460)
                     /label="SV40_enhancer"
                     /translation="MASLLKKTMPFRTTIEGTVNGHYFKCTGKGEGNPLEGTQEMKIE
                     VIEGGPLPFAFHILSTSCMYGSKAFIKYVSGIPDYFKQSLPEGFTWERTTTYEDGGFL
                     TAHQDTSLDGDCLVYKVKILGNNFPADGPVMQNKAGRWEPSTEIVYEVDGVLRGQSLM
                     ALECPGGRHLTCHLHTTYRSKKPASALKMPGFHFEDHRIEILEEVEKGKCYKQYEAAV
                     GRYCDAAPSKLGHN*"
     promoter        2257..2525
                     /label="SV40_promoter"
                     /translation="MASLLKKTMPFRTTIEGTVNGHYFKCTGKGEGNPLEGTQEMKIE
                     VIEGGPLPFAFHILSTSCMYGSKAFIKYVSGIPDYFKQSLPEGFTWERTTTYEDGGFL
                     TAHQDTSLDGDCLVYKVKILGNNFPADGPVMQNKAGRWEPSTEIVYEVDGVLRGQSLM
                     ALECPGGRHLTCHLHTTYRSKKPASALKMPGFHFEDHRIEILEEVEKGKCYKQYEAAV
                     GRYCDAAPSKLGHN*"
     rep_origin      2424..2501
                     /label="SV40_origin"
                     /translation="MASLLKKTMPFRTTIEGTVNGHYFKCTGKGEGNPLEGTQEMKIE
                     VIEGGPLPFAFHILSTSCMYGSKAFIKYVSGIPDYFKQSLPEGFTWERTTTYEDGGFL
                     TAHQDTSLDGDCLVYKVKILGNNFPADGPVMQNKAGRWEPSTEIVYEVDGVLRGQSLM
                     ALECPGGRHLTCHLHTTYRSKKPASALKMPGFHFEDHRIEILEEVEKGKCYKQYEAAV
                     GRYCDAAPSKLGHN*"
     misc_feature    2486..2505
                     /label="SV40pro_F_primer"
                     /translation="MASLLKKTMPFRTTIEGTVNGHYFKCTGKGEGNPLEGTQEMKIE
                     VIEGGPLPFAFHILSTSCMYGSKAFIKYVSGIPDYFKQSLPEGFTWERTTTYEDGGFL
                     TAHQDTSLDGDCLVYKVKILGNNFPADGPVMQNKAGRWEPSTEIVYEVDGVLRGQSLM
                     ALECPGGRHLTCHLHTTYRSKKPASALKMPGFHFEDHRIEILEEVEKGKCYKQYEAAV
                     GRYCDAAPSKLGHN*"
     CDS             2608..3402
                     /label="ORF frame 1"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGR
                     PVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDL
                     LSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRY
                     QDIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF*"
     gene            2611..3399
                     /label="NeoR/KanR"
                     /gene="NeoR/KanR"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGR
                     PVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDL
                     LSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRY
                     QDIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF*"
     CDS             complement(2917..3453)
                     /label="ORF frame 3"
                     /translation="MAGWASLGRSFRTPESRSEELVKKAIEGDALRIGSGDTVKHEEA
                     VSPFAAKLFSNITGSQRYVLIAVRHTQPATVDESRKAAIFHHDIRQAGIAMGHDEILA
                     VGHARLEPGEQFGWREPLMLFVQIILIDKTGFHPSTCSLDAMFRLVVEWAGSRIKRMQ
                     PPHCISHDGYFLGRSKVR*"
     terminator      3577..3846
                     /label="TK_PA_terminator"
                     /translation="MAGWASLGRSFRTPESRSEELVKKAIEGDALRIGSGDTVKHEEA
                     VSPFAAKLFSNITGSQRYVLIAVRHTQPATVDESRKAAIFHHDIRQAGIAMGHDEILA
                     VGHARLEPGEQFGWREPLMLFVQIILIDKTGFHPSTCSLDAMFRLVVEWAGSRIKRMQ
                     PPHCISHDGYFLGRSKVR*"
     rep_origin      3994..4613
                     /label="pBR322_origin"
                     /translation="MAGWASLGRSFRTPESRSEELVKKAIEGDALRIGSGDTVKHEEA
                     VSPFAAKLFSNITGSQRYVLIAVRHTQPATVDESRKAAIFHHDIRQAGIAMGHDEILA
                     VGHARLEPGEQFGWREPLMLFVQIILIDKTGFHPSTCSLDAMFRLVVEWAGSRIKRMQ
                     PPHCISHDGYFLGRSKVR*"
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 GGCCTCTTTG CTGAAGAAGA CCATGCCCTT CAGGACCACC
  721 ATCGAGGGCA CCGTGAACGG CCACTACTTC AAGTGCACCG GCAAGGGCGA GGGCAACCCC
  781 CTGGAGGGCA CCCAGGAGAT GAAGATCGAG GTGATCGAGG GCGGCCCCCT GCCCTTCGCC
  841 TTCCACATCC TGTCCACCTC CTGCATGTAC GGCTCCAAGG CCTTCATCAA GTACGTGTCC
  901 GGCATCCCCG ACTACTTCAA GCAGTCCCTC CCCGAGGGCT TCACCTGGGA GCGCACCACC
  961 ACCTACGAGG ACGGCGGCTT CCTGACCGCC CACCAGGACA CCTCCCTGGA CGGCGACTGC
 1021 CTGGTGTACA AGGTGAAGAT CCTGGGCAAC AACTTCCCCG CCGACGGCCC CGTGATGCAG
 1081 AACAAGGCCG GCCGCTGGGA GCCCTCCACC GAGATCGTGT ACGAGGTGGA CGGCGTGCTG
 1141 CGCGGCCAGT CCCTGATGGC CCTGGAGTGC CCCGGCGGTC GCCACCTGAC CTGCCACCTG
 1201 CACACCACCT ACCGCTCCAA GAAGCCCGCC TCCGCCCTGA AGATGCCCGG CTTCCACTTC
 1261 GAGGACCACC GCATCGAGAT CCTGGAGGAG GTGGAGAAGG GCAAGTGCTA CAAGCAGTAC
 1321 GAGGCCGCCG TGGGCCGCTA CTGCGACGCC GCCCCCTCCA AGCTGGGCCA CAACTGAAGC
 1381 GGCCGCGACT CTAGATCATA ATCAGCCATA CCACATTTGT AGAGGTTTTA CTTGCTTTAA
 1441 AAAACCTCCC ACACCTCCCC CTGAACCTGA AACATAAAAT GAATGCAATT GTTGTTGTTA
 1501 ACTTGTTTAT TGCAGCTTAT AATGGTTACA AATAAAGCAA TAGCATCACA AATTTCACAA
 1561 ATAAAGCATT TTTTTCACTG CATTCTAGTT GTGGTTTGTC CAAACTCATC AATGTATCTT
 1621 AAGGCGTAAA TTGTAAGCGT TAATATTTTG TTAAAATTCG CGTTAAATTT TTGTTAAATC
 1681 AGCTCATTTT TTAACCAATA GGCCGAAATC GGCAAAATCC CTTATAAATC AAAAGAATAG
 1741 ACCGAGATAG GGTTGAGTGT TGTTCCAGTT TGGAACAAGA GTCCACTATT AAAGAACGTG
 1801 GACTCCAACG TCAAAGGGCG AAAAACCGTC TATCAGGGCG ATGGCCCACT ACGTGAACCA
 1861 TCACCCTAAT CAAGTTTTTT GGGGTCGAGG TGCCGTAAAG CACTAAATCG GAACCCTAAA
 1921 GGGAGCCCCC GATTTAGAGC TTGACGGGGA AAGCCGGCGA ACGTGGCGAG AAAGGAAGGG
 1981 AAGAAAGCGA AAGGAGCGGG CGCTAGGGCG CTGGCAAGTG TAGCGGTCAC GCTGCGCGTA
 2041 ACCACCACAC CCGCCGCGCT TAATGCGCCG CTACAGGGCG CGTCAGGTGG CACTTTTCGG
 2101 GGAAATGTGC GCGGAACCCC TATTTGTTTA TTTTTCTAAA TACATTCAAA TATGTATCCG
 2161 CTCATGAGAC AATAACCCTG ATAAATGCTT CAATAATATT GAAAAAGGAA GAGTCCTGAG

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