| 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: | 高 |
| 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: | 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 |
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pEGFP-N1载体 pEGFPN1质粒基本信息
pEGFP-N1质粒图谱 pEGFPN1载体图谱和pEGFP-N1载体序列 pEGFPN1质粒序列多克隆位点信息
pEGFP-N1质粒 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).
pEGFP-N1质粒序列 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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pEGFP-N1质粒 pEGFPN1载体应用举例
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