- BioVector NTCC典型培養物保藏中心
- 聯系人：Dr.Xu, Biovector NTCC Inc.
電話：400-800-2947 工作QQ:1843439339 (微信同號)
郵件：Biovector@163.com
手機：18901268599
地址：北京
- 已注冊
| Clontech |
| pIREShyg3 |
| 哺乳動物細胞表達載體 |
| CMV |
| 高 |
| 多克隆位點，限制性內切酶 |
| 5752 bp |
| CMV-F: 5'-CGCAAATGGGCGGTAGGCGTG-3' |
| pIRES-R:5'-GCCCTAGATGCATGCTCG-3' |
| / |
| Ampicillin |
| Hygromycin |
Stably select with hygromycin |
| 631620 |
| 穩表達 Stable |
| 組成型 Constitutive |
| 非病毒 |
載體質粒圖譜和多克隆位點信息
載體簡介
Description
pIREShyg3 contains the internal ribosome entry site (IRES) of the encephalomyocarditis virus (ECMV), which permits the translation of two open reading frames from one messenger RNA (1–3). After selection with hygromycin B, nearly all surviving colonies will stably express the gene of interest, thus decreasing the need to screen large numbers of colonies to find functional clones. To select for cells that express high levels of the gene of interest, the selective pressure for antibiotic resistance was increased by shifting the hygromycin B phosphotransferase gene downstream to a less optimal position for translation as directed by the IRES sequence (3). By decreasing the level of expression of the antibiotic resistance marker, the selective pressure on the entire expression cassette is increased, resulting in selection for cells that express the entire transcript, including the gene of interest, at high levels. The expression cassette of pIREShyg3 contains the human cytomegalovirus (CMV) major immediate early promoter/enhancer followed by a multiple cloning site (MCS) that precedes stop codons in all three reading frames, a synthetic intron known to enhance the stability of the mRNA (4), the ECMV IRES followed by the hygromycin B phosphotransferase gene, and the polyadenylation signal from SV40. Ribosomes can enter the bicistronic mRNA at the 5' end to translate the gene of interest and at the ECMV IRES to translate the antibiotic resistance marker.
Use
When using the pIREShyg3 Vector, the antibiotic exerts selective pressure on the entire expression cassette; thus, a high dose of antibiotic will select for cells expressing a high level of the gene of interest. This selective pressure also ensures that the expression of the gene of interest will be stable over time in culture. Unless your expression experiments require a pure population of cells, you can use the pool of cells surviving selection instead of isolating and characterizing clonal cell lines. We recommend selecting mammalian cultures in 250–600 μg/ml Hygromycin B (Cat. No. 631309) depending on the cell line. Be sure to establish a kill curve for each cell line and each lot of Hygromycin B to determine optimal selection concentration.
載體序列
61 CCGCATAGTT AAGCCAGTAT CTGCTCCCTG CTTGTGTGTT GGAGGTCGCT GAGTAGTGCG
121 CGAGCAAAAT TTAAGCTACA ACAAGGCAAG GCTTGACCGA CAATTGCATG AAGAATCTGC
181 TTAGGGTTAG GCGTTTTGCG CTGCTTCGCG ATGTACGGGC CAGATATACG CGTTGACATT
241 GATTATTGAC TAGTTATTAA TAGTAATCAA TTACGGGGTC ATTAGTTCAT AGCCCATATA
301 TGGAGTTCCG CGTTACATAA CTTACGGTAA ATGGCCCGCC TGGCTGACCG CCCAACGACC
361 CCCGCCCATT GACGTCAATA ATGACGTATG TTCCCATAGT AACGCCAATA GGGACTTTCC
421 ATTGACGTCA ATGGGTGGAC TATTTACGGT AAACTGCCCA CTTGGCAGTA CATCAAGTGT
481 ATCATATGCC AAGTACGCCC CCTATTGACG TCAATGACGG TAAATGGCCC GCCTGGCATT
541 ATGCCCAGTA CATGACCTTA TGGGACTTTC CTACTTGGCA GTACATCTAC GTATTAGTCA
601 TCGCTATTAC CATGGTGATG CGGTTTTGGC AGTACATCAA TGGGCGTGGA TAGCGGTTTG
661 ACTCACGGGG ATTTCCAAGT CTCCACCCCA TTGACGTCAA TGGGAGTTTG TTTTGGCACC
721 AAAATCAACG GGACTTTCCA AAATGTCGTA ACAACTCCGC CCCATTGACG CAAATGGGCG
781 GTAGGCGTGT ACGGTGGGAG GTCTATATAA GCAGAGCTCT CTGGCTAACT AGAGAACCCA
841 CTGCTTACTG GCTTATCGAA ATTAATACGA CTCACTATAG GGAGACCCAA GCTTGGTACC
901 GAGCTCGGAT CTGTACAGGC CTTAAGCCGG CGCGCTAGCG CTTCGAAGGC GCCCGGGATC
961 CGATATCGTA CGTTAACATA GATAACTGAT CCAGTGTGCT GGAATTAATT CGCTGTCTGC
1021 GAGGGCCAGC TGTTGGGGTG AGTACTCCCT CTCAAAAGCG GGCATGACTT CTGCGCTAAG
1081 ATTGTCAGTT TCCAAAAACG AGGAGGATTT GATATTCACC TGGCCCGCGG TGATGCCTTT
1141 GAGGGTGGCC GCGTCCATCT GGTCAGAAAA GACAATCTTT TTGTTGTCAA GCTTGAGGTG
1201 TGGCAGGCTT GAGATCTGGC CATACACTTG AGTGACAATG ACATCCACTT TGCCTTTCTC
1261 TCCACAGGTG TCCACTCCCA GGTCCAACTG CAGGTCGAGC ATGCATCTAG GGCGGCCGCA
1321 CTAGAGGAAT TCCGCCCCTC TCCCTCCCCC CCCCCTAACG TTACTGGCCG AAGCCGCTTG
1381 GAATAAGGCC GGTGTGTGTT TGTCTATATG TTATTTTCCA CCATATTGCC GTCTTTTGGC
1441 AATGTGAGGG CCCGGAAACC TGGCCCTGTC TTCTTGACGA GCATTCCTAG GGGTCTTTCC
1501 CCTCTCGCCA AAGGAATGCA AGGTCTGTTG AATGTCGTGA AGGAAGCAGT TCCTCTGGAA
1561 GCTTCTTGAA GACAAACAAC GTCTGTAGCG ACCCTTTGCA GGCAGCGGAA CCCCCCACCT
1621 GGCGACAGGT GCCTCTGCGG CCAAAAGCCA CGTGTATAAG ATACACCTGC AAAGGCGGCA
1681 CAACCCCAGT GCCACGTTGT GAGTTGGATA GTTGTGGAAA GAGTCAAATG GCTCTCCTCA
1741 AGCGTAGTCA ACAAGGGGCT GAAGGATGCC CAGAAGGTAC CCCATTGTAT GGGAATCTGA
1801 TCTGGGGCCT CGGTGCACAT GCTTTACATG TGTTTAGTCG AGGTTAAAAA AACGTCTAGG
1861 CCCCCCGAAC CACGGGGACG TGGTTTTCCT TTGAAAAACA CGATGATAAG CTTGCCACAA
1921 CCCGTACCAA AGATGGATAG ATCCGGAAAG CCTGAACTCA CCGCGACGTC TGTCGAGAAG
1981 TTTCTGATCG AAAAGTTCGA CAGCGTCTCC GACCTGATGC AGCTCTCGGA GGGCGAAGAA
2041 TCTCGTGCTT TCAGCTTCGA TGTAGGAGGG CGTGGATATG TCCTGCGGGT AAATAGCTGC
2101 GCCGATGGTT TCTACAAAGA TCGTTATGTT TATCGGCACT TTGCATCGGC CGCGCTCCCG
2161 ATTCCGGAAG TGCTTGACAT TGGGGAATTC AGCGAGAGCC TGACCTATTG CATCTCCCGC
2221 CGTGCACAGG GTGTCACGTT GCAAGACCTG CCTGAAACCG AACTGCCCGC TGTTCTGCAG
2281 CCGGTCGCGG AGGCCATGGA TGCGATCGCT GCGGCCGATC TTAGCCAGAC GAGCGGGTTC
2341 GGCCCATTCG GACCGCAAGG AATCGGTCAA TACACTACAT GGCGTGATTT CATATGCGCG
2401 ATTGCTGATC CCCATGTGTA TCACTGGCAA ACTGTGATGG ACGACACCGT CAGTGCGTCC
2461 GTCGCGCAGG CTCTCGATGA GCTGATGCTT TGGGCCGAGG ACTGCCCCGA AGTCCGGCAC
2521 CTCGTGCACG CGGATTTCGG CTCCAACAAT GTCCTGACGG ACAATGGCCG CATAACAGCG
2581 GTCATTGACT GGAGCGAGGC GATGTTCGGG GATTCCCAAT ACGAGGTCGC CAACATCTTC
2641 TTCTGGAGGC CGTGGTTGGC TTGTATGGAG CAGCAGACGC GCTACTTCGA GCGGAGGCAT
2701 CCGGAGCTTG CAGGATCGCC GCGGCTCCGG GCGTATATGC TCCGCATTGG TCTTGACCAA
2761 CTCTATCAGA GCTTGGTTGA CGGCAATTTC GATGATGCAG CTTGGGCGCA GGGTCGATGC
2821 GACGCAATCG TCCGATCCGG AGCCGGGACT GTCGGGCGTA CACAAATCGC CCGCAGAAGC
2881 GCGGCCGTCT GGACCGATGG CTGTGTAGAA GTACTCGCCG ATAGTGGAAA CCGACGCCCC
2941 AGCACTCGTC CGAGGGCAAA GGAATAGAGT AGATGCCGAC CGAACAAGAG CTGATTTCGA
3001 GAACGCCTCA GCCAGCAACT CGCGCGAGCC TAGCAAGGCA AATGCGAGAG AACGGCCTTA
3061 CGCTTGGTGG CACAGTTCTC GTCCACAGTT CGCTAAGCTC GCTCGGCTGG GTCGCGGGAG
3121 GGCCGGTCGC AGTGATTCAG GCCCTTCTGG ATTGTGTTGG TCCCCAGGGC ACGATTGTCA
3181 TGCCCACGCA CTCGGGTGAT CTGACTGATC CCGCAGATTG GAGATCGCCG CCCGTGCCTG
3241 CCGATTGGGT GCAGATCTAG ATAACTGATC ATAATCAGCC ATACCACATT TGTAGAGGTT
3301 TTACTTGCTT TAAAAAACCT CCCACACCTC CCCCTGAACC TGAAACATAA AATGAATGCA
3361 ATTGTTGTTG TTAACTTGTT TATTGCAGCT TATAATGGTT ACAAATAAAG CAATAGCATC
3421 ACAAATTTCA CAAATAAAGC ATTTTTTTCA CTGCATTCTA GTTGTGGTTT GTCCAAACTC
3481 ATCAATGTAT CTTAACGCTC GAGTGCATTC TAGTTGTGGT TTGTCCAAAC TCATCAATGT
3541 ATCTTATCAT GTCTGTATAC CGTCGACCTC TAGCTAGAGC TTGGCGTAAT CATGGTCATA
3601 GCTGTTTCCT GTGTGAAATT GTTATCCGCT CACAATTCCA CACAACATAC GAGCCGGAAG
3661 CATAAAGTGT AAAGCCTGGG GTGCCTAATG AGTGAGCTAA CTCACATTAA TTGCGTTGCG
3721 CTCACTGCCC GCTTTCCAGT CGGGAAACCT GTCGTGCCAG CTGCATTAAT GAATCGGCCA
3781 ACGCGCGGGG AGAGGCGGTT TGCGTATTGG GCGCTCTTCC GCTTCCTCGC TCACTGACTC
3841 GCTGCGCTCG GTCGTTCGGC TGCGGCGAGC GGTATCAGCT CACTCAAAGG CGGTAATACG
3901 GTTATCCACA GAATCAGGGG ATAACGCAGG AAAGAACATG TGAGCAAAAG GCCAGCAAAA
3961 GGCCAGGAAC CGTAAAAAGG CCGCGTTGCT GGCGTTTTTC CATAGGCTCC GCCCCCCTGA
4021 CGAGCATCAC AAAAATCGAC GCTCAAGTCA GAGGTGGCGA AACCCGACAG GACTATAAAG
4081 ATACCAGGCG TTTCCCCCTG GAAGCTCCCT CGTGCGCTCT CCTGTTCCGA CCCTGCCGCT
4141 TACCGGATAC CTGTCCGCCT TTCTCCCTTC GGGAAGCGTG GCGCTTTCTC AATGCTCACG
4201 CTGTAGGTAT CTCAGTTCGG TGTAGGTCGT TCGCTCCAAG CTGGGCTGTG TGCACGAACC
4261 CCCCGTTCAG CCCGACCGCT GCGCCTTATC CGGTAACTAT CGTCTTGAGT CCAACCCGGT
4321 AAGACACGAC TTATCGCCAC TGGCAGCAGC CACTGGTAAC AGGATTAGCA GAGCGAGGTA
4381 TGTAGGCGGT GCTACAGAGT TCTTGAAGTG GTGGCCTAAC TACGGCTACA CTAGAAGGAC
4441 AGTATTTGGT ATCTGCGCTC TGCTGAAGCC AGTTACCTTC GGAAAAAGAG TTGGTAGCTC
4501 TTGATCCGGC AAACAAACCA CCGCTGGTAG CGGTGGTTTT TTTGTTTGCA AGCAGCAGAT
4561 TACGCGCAGA AAAAAAGGAT CTCAAGAAGA TCCTTTGATC TTTTCTACGG GGTCTGACGC
4621 TCAGTGGAAC GAAAACTCAC GTTAAGGGAT TTTGGTCATG AGATTATCAA AAAGGATCTT
4681 CACCTAGATC CTTTTAAATT AAAAATGAAG TTTTAAATCA ATCTAAAGTA TATATGAGTA
4741 AACTTGGTCT GACAGTTACC AATGCTTAAT CAGTGAGGCA CCTATCTCAG CGATCTGTCT
4801 ATTTCGTTCA TCCATAGTTG CCTGACTCCC CGTCGTGTAG ATAACTACGA TACGGGAGGG
4861 CTTACCATCT GGCCCCAGTG CTGCAATGAT ACCGCGAGAC CCACGCTCAC CGGCTCCAGA
4921 TTTATCAGCA ATAAACCAGC CAGCCGGAAG GGCCGAGCGC AGAAGTGGTC CTGCAACTTT
4981 ATCCGCCTCC ATCCAGTCTA TTAATTGTTG CCGGGAAGCT AGAGTAAGTA GTTCGCCAGT
5041 TAATAGTTTG CGCAACGTTG TTGCCATTGC TACAGGCATC GTGGTGTCAC GCTCGTCGTT
5101 TGGTATGGCT TCATTCAGCT CCGGTTCCCA ACGATCAAGG CGAGTTACAT GATCCCCCAT
5161 GTTGTGCAAA AAAGCGGTTA GCTCCTTCGG TCCTCCGATC GTTGTCAGAA GTAAGTTGGC
5221 CGCAGTGTTA TCACTCATGG TTATGGCAGC ACTGCATAAT TCTCTTACTG TCATGCCATC
5281 CGTAAGATGC TTTTCTGTGA CTGGTGAGTA CTCAACCAAG TCATTCTGAG AATAGTGTAT
5341 GCGGCGACCG AGTTGCTCTT GCCCGGCGTC AATACGGGAT AATACCGCGC CACATAGCAG
5401 AACTTTAAAA GTGCTCATCA TTGGAAAACG TTCTTCGGGG CGAAAACTCT CAAGGATCTT
5461 ACCGCTGTTG AGATCCAGTT CGATGTAACC CACTCGTGCA CCCAACTGAT CTTCAGCATC
5521 TTTTACTTTC ACCAGCGTTT CTGGGTGAGC AAAAACAGGA AGGCAAAATG CCGCAAAAAA
5581 GGGAATAAGG GCGACACGGA AATGTTGAAT ACTCATACTC TTCCTTTTTC AATATTATTG
5641 AAGCATTTAT CAGGGTTATT GTCTCATGAG CGGATACATA TTTGAATGTA TTTAGAAAAA
5701 TAAACAAATA GGGGTTCCGC GCACATTTCC CCGAAAAGTG CCACCTGACG TC
//
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電郵Biovector@163.com
電話18901268599
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