| 出品公司: | Clontech |
|---|---|
| 载体名称: | pAcGFP1-Lam |
| 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: | 5462 bp |
| 5' 测序引物及序列: | -- |
| 3' 测序引物及序列: | -- |
| 载体标签: | AcGFP1 (N-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pAcGFP1-Lam组成型表达N端AcGFP融合蛋白LaminA/C； pAcGFP1-Lam是MatchMaker系统载体； AcGFP1与GFP相比，密码子经过了优化，更适合在哺乳动物细胞中高水平表达，同时也增强了亮度。 |
| 产品目录号: | 630458 |
| 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 |
买家导航
pAcGFP1-Lam载体质粒基本信息
pAcGFP1-Lam质粒图谱载体图谱和pAcGFP1-Lam载体序列质粒序列多克隆位点信息
pAcGFP1-Lam质粒载体简介
pAcGFP1-Lam载体描述
pAcGFP1-Lamin encodes a green fluorescent protein (GFP) from Aequorea coerulescens (excitation maximum = 475 nm; emission maximum = 505 nm) and the gene encoding the human nuclear lamin (1). SV40 polyadenylation signals downstream of the AcGFP1-Lamin fusion direct proper processing of the 3’end of the AcGFP1-Lamin mRNA. AcGFP1 contains silent mutations that create an open reading frame comprised almost entirely of optimized human codons. These changes increase the translational efficiency of the AcGFP1 mRNA and consequently the expression of AcGFP1 in mammalian and plant cells. The vector backbone also contains an SV40 origin for replication in any mammalian cell line that expresses 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 to be selected using G418. A bacterial promoter upstream of this cassette drives expression of the gene encoding kanamycin resistance in E. coli. The pAcGFP1-Lamin backbone also provides a pUC origin of replication for propagation in E. coli and an f1 origin for single-stranded DNA production.
The pAcGFP1-Lamin is provided in the Matchmaker Chemiluminescent Co-IP Vector Set (Cat. No. 630458) as a negative bait control construct. In mammalian cells, this vector expresses the AcGFP1-Lamin fusion protein—the AcGFP1 serves as a tag, through which a polyclonal antibody against AcGFP1 can be used for immunoprecipitation, while lamin serves as the bait for the capture of interacting proteins via coimmunoprecipitation. Because lamin does not interact with SV40 large T antigen (2), in lysates of cells cotransfected with pAcGFP1-Lamin and pProLabel-T, there is no ProLabel activity detected in the Co-IP of this control for noninteracting pairs of proteins. The fluorescence from AcGFP1-Lamin fusion protein also allows for easy detection of fusion expression and localization in vivo, as well as offering a mode for determining transfection efficiency.
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) in E. coli hosts.
E. coli replication origin: pUC
Copy number: ~500
Plasmid incompatibility group: pMB1/ColE1
References
1. Sturman, N. et al. (1998) J. Stuct. Biol. 122(1–2) 42–66.
2. Matchmaker Co-IP Kit (April 1999) Clontechniques XIV(2): 14–15.
3. Gorman, C. (1985) In DNA Cloning: A Practical Approach, Vol. II. Ed. D.M. Glover. (IRL Press, Oxford, UK) pp. 143–190.
pAcGFP1-Lam质粒序列载体序列
LOCUS pAcGFP1-Lam 5462 bp DNA circular 1-FEB-2010
COMMENT Created by Clontech Laboratories Inc. http://www.clontech.com
COMMENT Cat. No. 630458
FEATURES Location/Qualifiers
misc_feature 1..589
/label=CMV\IE\Promoter
misc_feature 613..1329
/label=AcGFP1
misc_feature 1426..1920
/label=Lamin\A/C
misc_feature 2281..2331
/label=SV40\early\poly\signals
misc_feature 2378..2833
/label=f1\origin
misc_feature 3174..3309
/label=SV40\origin
misc_feature 3358..4152
/label=Kan(R)/Neo(R)
misc_feature 4388..4406
/label=HSV\TK\polyA\signals
misc_feature 4737..5380
/label=pUC\origin
BASE COUNT 1328 a 1439 c 1472 g 1223 t
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 ccggtcgcca ccatggtgag caagggcgcc gagctgttca ccggcatcgt gcccatcctg
661 atcgagctga atggcgatgt gaatggccac aagttcagcg tgagcggcga gggcgagggc
721 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcctgtg
781 ccctggccca ccctggtgac caccctgagc tacggcgtgc agtgcttctc acgctacccc
841 gatcacatga agcagcacga cttcttcaag agcgccatgc ctgagggcta catccaggag
901 cgcaccatct tcttcgagga tgacggcaac tacaagtcgc gcgccgaggt gaagttcgag
961 ggcgataccc tggtgaatcg catcgagctg accggcaccg atttcaagga ggatggcaac
1021 atcctgggca ataagatgga gtacaactac aacgcccaca atgtgtacat catgaccgac
1081 aaggccaaga atggcatcaa ggtgaacttc aagatccgcc acaacatcga ggatggcagc
1141 gtgcagctgg ccgaccacta ccagcagaat acccccatcg gcgatggccc tgtgctgctg
1201 cccgataacc actacctgtc cacccagagc gccctgtcca aggaccccaa cgagaagcgc
1261 gatcacatga tctacttcgg cttcgtgacc gccgccgcca tcacccacgg catggatgag
1321 ctgtacaagt ccggacttaa ggcctctgtc gacgccgccg aggagcagaa gctgatctca
1381 gaggaggacc tgcatatggc catggaggcc gaattcccgg ggatctctga agaggtggtc
1441 agccgcgagg tgtccggcat caaggccgcc tacgaggccg agctcgggga tgcccgcaag
1501 acccttgact cagtagccaa ggagcgcgcc cgcctgcagc tggagctgag caaagtgcgt
1561 gaggagttta aggagctgaa agcgcgcaat accaagaagg agggtgacct gatagctgct
1621 caggctcggc tgaaggacct ggaggctctg ctgaactcca aggaggccgc actgagcact
1681 gctctcagtg agaagcgcac gctggagggc gagctgcatg atctgcgggg ccaggtggcc
1741 aagcttgagg cagccctagg tgaggccaag aagcaacttc aggatgagat gctgcggcgg
1801 gtggatgctg agaacaggct gcagaccatg aaggaggaac tggacttcca gaagaacatc
1861 tacagtgagg agctgcgtga gaccaagcgc cgtcatgaga cccgactggt ggagattgac
1921 aggatccgtc gacctgcagc ggccgcataa ctagcataac cccttggggc ctctaaacgg
1981 gtcttgaggg gttttttgcg cgcttgcagc caagctaatt ccgggcgaat ttcttatgat
2041 ttatgatttt tattattaaa taagttataa aaaaaataag tgtatacaaa ttttaaagtg
2101 acaagcttgc gaattctggc ggatccaccg gatctagata actgatcata atcagccata
2161 ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc ctgaacctga
2221 aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat aatggttaca
2281 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
2341 gtggtttgtc caaactcatc aatgtatctt aacgcgtaaa ttgtaagcgt taatattttg
2401 ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc
2461 ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt
2521 tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc
2581 tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg
2641 tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga
2701 aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg
2761 ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg
2821 ctacagggcg cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
2881 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
2941 caataatatt gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg aatgtgtgtc
3001 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
3061 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
3121 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
3181 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
3241 atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt
3301 ttggaggcct aggcttttgc aaagatcgat caagagacag gatgaggatc gtttcgcatg
3361 attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc
3421 tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg
3481 caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa
3541 gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc
3601 gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat
3661 ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg
3721 cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc
3781 gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag
3841 catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgagcat gcccgacggc
3901 gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc
3961 cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata
4021 gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc
4081 gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac
4141 gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc
4201 catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
4261 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
4321 accctagggg gaggctaact gaaacacgga aggagacaat accggaagga acccgcgcta
4381 tgacggcaat aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt cataaacgcg
4441 gggttcggtc ccagggctgg cactctgtcg ataccccacc gagaccccat tggggccaat
4501 acgcccgcgt ttcttccttt tccccacccc accccccaag ttcgggtgaa ggcccagggc
4561 tcgcagccaa cgtcggggcg gcaggccctg ccatagcctc aggttactca tatatacttt
4621 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
4681 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
4741 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
4801 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
4861 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
4921 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
4981 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
5041 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
5101 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
5161 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
5221 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
5281 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
5341 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
5401 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgccatgc
5461 at
//
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