| 出品公司: | Clontech |
|---|---|
| 载体名称: | pRetroX-Tight-Hyg-Luc |
| 质粒类型: | 逆病毒载体；四环素调控载体；荧光素酶报告载体 |
| 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: | Tight |
| 载体大小: | 9054 bp |
| 5' 测序引物及序列: | 5’-ATCTGAGGCCCTTTCGTCTTCACT-3’ |
| 3' 测序引物及序列: | -- |
| 载体标签: | 无 |
| 载体抗性: | 氨苄青霉素 |
| 筛选标记: | 潮霉素（Hygromycin） |
| 克隆菌株: | DH5α |
| 宿主细胞（系）: | 常规细胞系（293、CV-1、CHO等） |
| 备注: | 逆病毒载体pRetroX-Tight-Hyg-Luc是四环素诱导载体，作为pRetroX-Tight-Hyg载体的阳性对照。 |
| 产品目录号: | 631034 |
| 稳定性: | 稳表达 |
| 组成型/诱导型: | 诱导型 |
| 病毒/非病毒: | 逆转录病毒 |
买家导航
载体基本信息
载体图谱质粒图谱和多克隆位点信息
载体简介
pRetroX-Tight-Hyg-Luc载体描述
pRetroX-Tight-Hyg-Luc is a retroviral control vector (supplied with pRetroX-Tight-Hyg) that expresses fi refl y luciferase under the control of PTight, a modifi ed Tet-responsive promoter. PTight consists of seven direct repeats of a 36 bp regulatory sequence that contains the 19 bp tet operator sequence (tetO), and a modifi ed minimal CMV promoter (1). This vector is designed to be used as a control vector with Clontech's Advanced Inducible Expression Systems. These systems provide ready access to the inducible gene expression strategy of Gossen & Bujard, and incorporate major improvements described by Urlinger, et al. (2-6).
pRetroX-Tight-Hyg-Luc is optimized to eliminate promoter interference through LTR selfinactivation. The hybrid 5’ LTR consists of the cytomegalovirus (CMV) type I enhancer and the mouse sarcoma virus (MSV) promoter. This promoter drives high levels of viral genome transcription in HEK 293-based packaging cell lines due, in part, to the presence of adenoviral E1A (7-8) in these cells. The self-inactivating feature of the vector is provided by a deletion in the 3’ LTR enhancer region (U3). During reverse transcription of the retroviral RNA, a copy of the inactivated 3’ LTR U3 region replaces the corresponding region of the 5’ LTR, resultingin the inactivation of the 5’ LTR CMV enhancer sequence. This mechanism can reduce the phenomenon known as promoter interference (9-10) and allow more effi cient expression. In
this way, pRetroX-Tight-Hyg-Luc supports high viral titers, yet eliminates potential downstream interference between the inducible PTight promoter and its adjacent viral LTR.
pRetroX-Tight-Hyg-Luc contains all of the necessary viral RNA processing elements; these include the 5’ and 3’ LTRs, the packaging signal (Ψ+), and the tRNA primer binding site. For safety reasons, however, the vector lacks the structural genes (gag, pol, and env) necessary for retroviral particle formation and replication. pRetroX-Tight-Hyg-Luc contains a hygromycin resistance gene (Hygr) under the control of the murine phosphoglycerate kinase (PGK) promoter (PPGK) for the selection of stable transfectants. In addition, the vector contains a ColE1 origin of replication and an E. coli Ampr gene for propagation and selection in bacteria.
pRetroX-Tight-Hyg-Luc can be used as a control vector for either plasmid or retroviral expression vectors. When used as a control for retroviral expression vectors, it must be transfected into a packaging cell line, such as GP2-293; we recommend the Retro-X Universal Packaging System (Cat. No. 631530). Packaging cell lines allow you to produce infectious, replication-incompetent retroviral particles. These retroviral particles can infect a wide range of target cells, but they cannot replicate within these cells due to the absence of viral structural genes. The separate introduction and integration of the structural genes into the packaging cell line minimizes the chance of producing replication-competent virus due to recombination events during cell proliferation.
Propagation in E. coli
Suitable host strains: DH5α and other general purpose strains.
Selectable marker: plasmid confers resistance to ampicillin (100 μg/ml) in E. coli hosts.
E. coli replication origin: ColE1
Copy number: high
Notes:
The attached sequence fi le has been compiled from information in the sequence databases, published literature, and other sources, together with partial sequences obtained by Clontech. This vector has not been completely sequence.
The viral supernatants produced by this retroviral vector could contain potentially hazardous recombinant virus. Due caution must be exercised in the production and handling of recombinant retrovirus. Appropriate NIH, regional, and institutional guidelines apply.
载体序列
LOCUS pRetroX-Tight-Hyg-Luc 9052 bp DNA circular 10-NOV-2009
COMMENT Created by Clontech Laboratories Inc. http://www.clontech.com
COMMENT Cat. No. 631034
FEATURES Location/Qualifiers
promoter 3602..4105
/label=PGK\Promoter
misc_feature 4123..5148
/label=Hyg(R)
LTR 5748..6184
/label=3'\MMLV\LTR\(del)
CDS complement(7830..8690)
/label=Amp(R)
promoter 1602..1917
/label=Tight\Promoter
LTR 1..727
/label=5'\LTR\(CMV/MSV)
misc_feature 758..1567
/label=Packaging\Signal
promoter 6463..6730
/label=SV40\Promoter
rep_origin 6684..6749
/label=SV40\origin
rep_origin 7071..7071
/label=pUC\origin
misc_feature 1934..3586
/label=Luciferase
BASE COUNT 2184 a 2335 c 2332 g 2201 t
ORIGIN
1 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
61 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
121 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
181 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc
241 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
301 ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac
361 ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc
421 aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc
481 gtgtacggtg ggaggtctat ataagcagag ctcaataaaa gagcccacaa cccctcactc
541 ggcgcgccag tcttccgata gactgcgtcg cccgggtacc cgtattccca ataaagcctc
601 ttgctgtttg catccgaatc gtggtctcgc tgttccttgg gagggtctcc tctgagtgat
661 tgactaccca cgacgggggt ctttcatttg ggggctcgtc cgggatttgg agacccctgc
721 ccagggacca ccgacccacc accgggaggt aagctggcca gcaacttatc tgtgtctgtc
781 cgattgtcta gtgtctatgt ttgatgttat gcgcctgcgt ctgtactagt tagctaacta
841 gctctgtatc tggcggaccc gtggtggaac tgacgagttc tgaacacccg gccgcaaccc
901 tgggagacgt cccagggact ttgggggccg tttttgtggc ccgacctgag gaagggagtc
961 gatgtggaat ccgaccccgt caggatatgt ggttctggta ggagacgaga acctaaaaca
1021 gttcccgcct ccgtctgaat ttttgctttc ggtttggaac cgaagccgcg cgtcttgtct
1081 gctgcagcgc tgcagcatcg ttctgtgttg tctctgtctg actgtgtttc tgtatttgtc
1141 tgaaaattag ggccagactg ttaccactcc cttaagtttg accttaggtc actggaaaga
1201 tgtcgagcgg atcgctcaca accagtcggt agatgtcaag aagagacgtt gggttacctt
1261 ctgctctgca gaatggccaa cctttaacgt cggatggccg cgagacggca cctttaaccg
1321 agacctcatc acccaggtta agatcaaggt cttttcacct ggcccgcatg gacacccaga
1381 ccaggtcccc tacatcgtga cctgggaagc cttggctttt gacccccctc cctgggtcaa
1441 gccctttgta caccctaagc ctccgcctcc tcttcctcca tccgccccgt ctctccccct
1501 tgaacctcct cgttcgaccc cgcctcgatc ctccctttat ccagccctca ctccttctct
1561 aggcgccgga attgaagatc tgaggccctt tcgtcttcac tcgagtttac tccctatcag
1621 tgatagagaa cgtatgtcga gtttactccc tatcagtgat agagaacgat gtcgagttta
1681 ctccctatca gtgatagaga acgtatgtcg agtttactcc ctatcagtga tagagaacgt
1741 atgtcgagtt tactccctat cagtgataga gaacgtatgt cgagtttatc cctatcagtg
1801 atagagaacg tatgtcgagt ttactcccta tcagtgatag agaacgtatg tcgaggtagg
1861 cgtgtacggt gggaggccta tataagcaga gctcgtttag tgaaccgtca gatcgcctgg
1921 agaaggatcc accatggaag acgccaaaaa cataaagaaa ggcccggcgc cattctatcc
1981 gctagaggat ggaaccgctg gagagcaact gcataaggct atgaagagat acgccctggt
2041 tcctggaaca attgctttta cagatgcaca tatcgaggtg aacatcacgt acgcggaata
2101 cttcgaaatg tccgttcggt tggcagaagc tatgaaacga tatgggctga atacaaatca
2161 cagaatcgtc gtatgcagtg aaaactctct tcaattcttt atgccggtgt tgggcgcgtt
2221 atttatcgga gttgcagttg cgcccgcgaa cgacatttat aatgaacgtg aattgctcaa
2281 cagtatgaac atttcgcagc ctaccgtagt gtttgtttcc aaaaaggggt tgcaaaaaat
2341 tttgaacgtg caaaaaaaat taccaataat ccagaaaatt attatcatgg attctaaaac
2401 ggattaccag ggatttcagt cgatgtacac gttcgtcaca tctcatctac ctcccggttt
2461 taatgaatac gattttgtac cagagtcctt tgatcgtgac aaaacaattg cactgataat
2521 gaactcctct ggatctactg ggttacctaa gggtgtggcc cttccgcata gaactgcctg
2581 cgtcagattc tcgcatgcca gagatcctat ttttggcaat caaatcattc cggatactgc
2641 gattttaagt gttgttccat tccatcacgg ttttggaatg tttactacac tcggatattt
2701 gatatgtgga tttcgagtcg tcttaatgta tagatttgaa gaagagctgt ttttacgatc
2761 ccttcaggat tacaaaattc aaagtgcgtt gctagtacca accctatttt cattcttcgc
2821 caaaagcact ctgattgaca aatacgattt atctaattta cacgaaattg cttctggggg
2881 cgcacctctt tcgaaagaag tcggggaagc ggttgcaaaa cgcttccatc ttccagggat
2941 acgacaagga tatgggctca ctgagactac atcagctatt ctgattacac ccgaggggga
3001 tgataaaccg ggcgcggtcg gtaaagttgt tccatttttt gaagcgaagg ttgtggatct
3061 ggataccggg aaaacgctgg gcgttaatca gagaggcgaa ttatgtgtca gaggacctat
3121 gattatgtcc ggttatgtaa acaatccgga agcgaccaac gccttgattg acaaggatgg
3181 atggctacat tctggagaca tagcttactg ggacgaagac gaacacttct tcatagttga
3241 ccgcttgaag tctttaatta aatacaaagg ataccaggtg gcccccgctg aattggagtc
3301 gatattgtta caacacccca acatcttcga cgcgggcgtg gcaggtcttc ccgacgatga
3361 cgccggtgaa cttcccgccg ccgttgttgt tttggagcac ggaaagacga tgacggaaaa
3421 agagatcgtg gattacgtcg ccagtcaagt aacaaccgcg aaaaagttgc gcggaggagt
3481 tgtgtttgtg gacgaagtac cgaaaggtct taccggaaaa ctcgacgcaa gaaaaatcag
3541 agagatcctc ataaaggcca agaagggcgg aaagtccaaa ttgtaaaata cgcgtgaatt
3601 ctaccgggta ggggaggcgc ttttcccaag gcagtctgga gcatgcgctt tagcagcccc
3661 gctgggcact tggcgctaca caagtggcct ctggcctcgc acacattcca catccaccgg
3721 taggcgccaa ccggctccgt tctttggtgg ccccttcgcg ccaccttcta ctcctcccct
3781 agtcaggaag ttcccccccg ccccgcagct cgcgtcgtgc aggacgtgac aaatggaagt
3841 agcacgtctc actagtctcg tgcagatgga cagcaccgct gagcaatgga agcgggtagg
3901 cctttggggc agcggccaat agcagctttg ctccttcgct ttctgggctc agaggctggg
3961 aaggggtggg tccgggggcg ggctcagggg cgggctcagg ggcggggcgg gcgcccgaag
4021 gtcctccgga ggcccggcat tctgcacgct tcaaaagcgc acgtctgccg cgctgttctc
4081 ctcttcctca tctccgggcc tttcgacctg catcccgcca ccatgaaaaa gcctgaactc
4141 accgcgacgt ctgtcgagaa gtttctgatc gaaaagttcg acagcgtctc cgacctgatg
4201 cagctctcgg agggcgaaga atctcgtgct ttcagcttcg atgtaggagg gcgtggatat
4261 gtcctgcggg taaatagctg cgccgatggt ttctacaaag atcgttatgt ttatcggcac
4321 tttgcatcgg ccgcgctccc gattccggaa gtgcttgaca ttggggagtt cagcgagagc
4381 ctgacctatt gcatctcccg ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc
4441 gaactgcccg ctgttctgca gccggtcgcg gaggccatgg atgcgatcgc tgcggccgat
4501 cttagccaga cgagcgggtt cggcccattc ggaccgcaag gaatcggtca atacactaca
4561 tggcgtgatt tcatatgcgc gattgctgat ccccatgtgt atcactggca aactgtgatg
4621 gacgacaccg tcagtgcgtc cgtcgcgcag gctctcgatg agctgatgct ttgggccgag
4681 gactgccccg aagtccggca cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg
4741 gacaatggcc gcataacagc ggtcattgac tggagcgagg cgatgttcgg ggattcccaa
4801 tacgaggtcg ccaacatctt cttctggagg ccgtggttgg cttgtatgga gcagcagacg
4861 cgctacttcg agcggaggca tccggagctt gcaggatcgc cgcggctccg ggcgtatatg
4921 ctccgcattg gtcttgacca actctatcag agcttggttg acggcaattt cgatgatgca
4981 gcttgggcgc agggtcgatg cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt
5041 acacaaatcg cccgcagaag cgcggccgtc tggaccgatg gctgtgtaga agtactcgcc
5101 gatagtggaa accgacgccc cagcactcgt ccgagggcaa aggaatagag tagatgccga
5161 ccgaacaaga gctgatttcg agaacgcctc agccagcaac tcgcgcgagc ctagcaaggc
5221 aaatgcgaga gaacggcctt acgcttggtg gcacagttct cgtccacagt tcgctaagct
5281 cgctcggctg ggtcgcggga gggccggtcg cagtgattca ggcccttctg gattgtgttg
5341 gtccccaggg cacgattgtc atgcccacgc actcgggtga tctgactgat cccgcagatt
5401 ggagatcgcc gcccgtgcct gccgattggg tgcagatccg tcgacctgca gccaagctta
5461 tcgataaaat aaaagatttt atttagtctc cagaaaaagg ggggaatgaa agaccccacc
5521 tgtaggtttg gcaagctagc ttaagtaacg ccattttgca aggcatggaa aatacataac
5581 tgagaataga gaagttcaga tcaaggttag gaacagagag acagcagaat atgggccaaa
5641 caggatatca gtggtccagg ctctagtttt gactcaacaa tatcaccagc tgaagcctat
5701 agagtacgag ccatagataa aataaaagat tttatttagt ctccagaaaa aggggggaat
5761 gaaagacccc acctgtaggt ttggcaagct agcttaagta acgccatttt gcaaggcatg
5821 gaaaaataca taactgagaa tagagaagtt cagatcaagg tcaggaacag atggaacagg
5881 gtcgacccta gagaaccatc agatgtttcc agggtgcccc aaggacctga aatgaccctg
5941 tgccttattt gaactaacca atcagttcgc ttctcgcttc tgttcgcgcg cttctgctcc
6001 ccgagctcaa taaaagagcc cacaacccct cactcggggc gccagtcctc cgattgactg
6061 agtcgcccgg gtacccgtgt atccaataaa ccctcttgca gttgcatccg acttgtggtc
6121 tcgctgttcc ttgggagggt ctcctctgag tgattgacta cccgtcagcg ggggtctttc
6181 atttgggggc tcgtccggga tcgggagacc cctgcccagg gaccaccgac ccaccaccgg
6241 gaggtaagct ggctgcctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca
6301 gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca
6361 gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt cacgtagcga
6421 tagcggagtg tagatccggc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
6481 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
6541 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
6601 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
6661 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc
6721 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
6781 gcttactggc ttaactatgc ggcatcagag cagattgtac tgagagtgca ccatatgcgg
6841 tgtgaaatac cgcacagatg cgtaaggaga aaataccgca tcaggcgctc ttccgcttcc
6901 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
6961 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
7021 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
7081 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
7141 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
7201 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
7261 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
7321 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
7381 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
7441 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
7501 tacactagaa ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
7561 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
7621 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
7681 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
7741 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
7801 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
7861 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
7921 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
7981 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
8041 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
8101 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctgcagg catcgtggtg
8161 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
8221 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
8281 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
8341 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
8401 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaacacg ggataatacc
8461 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
8521 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
8581 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
8641 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
8701 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
8761 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
8821 gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg
8881 ccctttcgtc ttcaagaatt agcttggcca ttgcatacgt tgtatccata tcataatatg
8941 tacatttata ttggctcatg tccaacatta ccgccatgtt gacattgatt attgactagt
9001 tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga gt
//
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