pEGFP-N1

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pEGFPN1载体基本信息

出品公司: Clontech
别名: pEGFP-N1, pEGFP N1
质粒类型: 哺乳动物细胞表达载体
启动子: CMV
表达水平:
克隆方法: 多克隆位点,限制性内切酶
载体大小: 4733bp
5' 测序引物: CMV-F
5' 测序引物序列:   5'-CGCAAATGGGCGGTAGGCGTG-3'
3' 测序引物: EGFP-N
3' 测序引物序列:   5'-CGTCGCCGTCCAGCTCGACCAG-3'
载体标签: C-EGFP
载体抗性: 卡那
筛选标记: Neomycin_1.html' target='_blank'>新霉素 (G418)
备注:
Clontech公司已经终止了该载体的生产和销售.
产品目录号: 6085-1
稳定性: 稳表达
组成型: 组成型
病毒/非病毒: 非病毒

pEGFPN1载体质粒图谱和多克隆位点信息

pEGFP-N1载体图谱和多克隆位点

pEGFPN1载体简介

pEGFP-N1 encodes a red-shifted variant of wild-type GFP (1–3) which has been optimized for brighter fluorescence and higher expression in mammalian cells. (Excitation maximum = 488 nm;emission maximum = 507 nm.) pEGFP-N1 encodes the GFPmut1 variant (4) which contains the double-amino-acid substitution of Phe-64 to Leu and Ser-65 to Thr. The coding sequence of the EGFP gene contains more than 190 silent base changes which correspond to human codon-usage preferences (5). Sequences flanking EGFP have been converted to a Kozak consensus translation initiation site (6) to further increase the translation efficiency in eukaryotic cells. The MCS in pEGFP-N1 is between the immediate early promoter of CMV (PCMV IE) and the EGFP coding sequences. Genes cloned into the MCS will be expressed as fusions to the N-terminus of EGFP if they are in the same reading frame as EGFP and there are no intervening stop codons. SV40polyadenylation signals downstream of the EGFP gene direct proper processing of the 3' end of the EGFP mRNA. The vector backbone also contains an SV40 origin for replication in mammalian cells expressing the SV40 T antigen. A neomycin-resistance cassette (Neor), consisting 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, allows stably transfected eukaryotic cells tobe selected using G418. A bacterial promoter upstream of this cassette expresses kanamycin resistance in E. coli. The pEGFP-N1 backbone also provides a pUC origin of replication for propagation in E. coli and an f1 origin for single-stranded DNA production.

Use:
Fusions to the N terminus of EGFP retain the fluorescent properties of the native protein allowing the localization of the fusion protein in vivo . The target gene should be cloned into pEGFP-N1 so that it is in frame with the EGFP coding sequences, with no intervening in-frame stop codons. The inserted gene should include the initiating ATG codon. The recombinant EGFP vector can be transfected into mammalian cells using any standard transfection method. If required, stable transformants can be selected using G418 (7). pEGFP-N1 can also be used simply to express EGFP in a cell line of interest (e.g., as a transfection marker).

pEGFPN1载体序列

LOCUS       pEGFP-N1    4733 bp     DNA Circular    SYN
DEFINITION  pEGFP-N1
ACCESSION   
KEYWORDS    
SOURCE      
  ORGANISM  other sequences; artificial sequences; vectors.
COMMENT     This file is created by Vector NTI
            http://www.biofeng.com/
COMMENT     ORIGDB|GenBank
COMMENT     VNTAUTHORNAME|biofeng.com|
FEATURES             Location/Qualifiers
     source          1..4733
                     /organism="pEGFP-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..1398
                     /label="ORF frame 1"
     gene            682..1395
                     /label="EGFP"
                     /gene="EGFP"
     misc_feature    complement(724..745)
                     /label="EGFP_N_primer"
     misc_feature    862..891
                     /label="Y66 (EGFP)"
     misc_feature    1332..1353
                     /label="EGFP_C_primer"
     misc_feature    1612..1631
                     /label="EBV_rev_primer"
     rep_origin      complement(1781..2087)
                     /label="f1_origin"
     promoter        2166..2194
                     /label="AmpR_promoter"
     misc_feature    complement(2260..2280)
                     /label="pBABE_3_primer"
     misc_feature    complement(2266..2481)
                     /label="SV40_enhancer"
     promoter        2278..2546
                     /label="SV40_promoter"
     rep_origin      2445..2522
                     /label="SV40_origin"
     misc_feature    2507..2526
                     /label="SV40pro_F_primer"
     CDS             2629..3423
                     /label="ORF frame 1"
     gene            2632..3420
                     /label="NeoR/KanR"
                     /gene="NeoR/KanR"
     CDS             complement(2938..3474)
                     /label="ORF frame 3"
     terminator      3598..3867
                     /label="TK_PA_terminator"
     rep_origin      4015..4634
                     /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 ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc
  721 atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc
  781 gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg
  841 cccgtgccct ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc
  901 taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc
  961 caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag
 1021 ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac
 1081 ggcaacatcc tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg
 1141 gccgacaagc agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac
 1201 ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg
 1261 ctgctgcccg acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag
 1321 aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg
 1381 gacgagctgt acaagtaaag cggccgcgac tctagatcat aatcagccat accacatttg
 1441 tagaggtttt acttgcttta aaaaacctcc cacacctccc cctgaacctg aaacataaaa
 1501 tgaatgcaat tgttgttgtt aacttgttta ttgcagctta taatggttac aaataaagca
 1561 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt
 1621 ccaaactcat caatgtatct taaggcgtaa attgtaagcg ttaatatttt gttaaaattc
 1681 gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc
 1741 ccttataaat caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag
 1801 agtccactat taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc
 1861 gatggcccac tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa
 1921 gcactaaatc ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg
 1981 aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt
 2041 gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc
 2101 gcgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
 2161 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
 2221 tgaaaaagga agagtcctga ggcggaaaga accagctgtg gaatgtgtgt cagttagggt
 2281 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
 2341 cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
 2401 atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc
 2461 cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg
 2521 ccgaggccgc ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc
 2581 taggcttttg caaagatcga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa
 2641 gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg
 2701 gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc
 2761 ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca agacgaggca
 2821 gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc
 2881 actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca
 2941 tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat
 3001 acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca
 3061 cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg
 3121 ctcgcgccag ccgaactgtt cgccaggctc aaggcgagca tgcccgacgg cgaggatctc
 3181 gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg ccgcttttct
 3241 ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat agcgttggct
 3301 acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct cgtgctttac
 3361 ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga cgagttcttc
 3421 tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg ccatcacgag
 3481 atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg
 3541 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccctaggg
 3601 ggaggctaac tgaaacacgg aaggagacaa taccggaagg aacccgcgct atgacggcaa
 3661 taaaaagaca gaataaaacg cacggtgttg ggtcgtttgt tcataaacgc ggggttcggt
 3721 cccagggctg gcactctgtc gataccccac cgagacccca ttggggccaa tacgcccgcg
 3781 tttcttcctt ttccccaccc caccccccaa gttcgggtga aggcccaggg ctcgcagcca
 3841 acgtcggggc ggcaggccct gccatagcct caggttactc atatatactt tagattgatt
 3901 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
 3961 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
 4021 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
 4081 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
 4141 taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag
 4201 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
 4261 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
 4321 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
 4381 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
 4441 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
 4501 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
 4561 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
 4621 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
 4681 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccatg cat

//

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