| 质粒名称: | pGL4.22-PGK1-HRE::dLUC |
|---|---|
| 出品公司: | Addgene |
| 目录编号: | 128095 |
| 存储实验室: | Chi Van Dang |
|
载体骨架基本信息 |
|
| 载体名称: | pGL4.22 |
| 载体出品公司: | Promega |
| 空载体大小: | 5566 bp |
| 完整质粒大小: | 5647 bp |
| 载体类型: | 哺乳细胞表达, Luciferase |
| 筛选标记: | Puromycin |
| 载体抗性: | 氨苄霉素 |
| 生长温度: | 37 ℃ |
| 宿主菌: | DH5a |
| 拷贝数: | 未知 |
|
插入基因信息 |
|
| 插入基因名称: | HIF response element (x3)、phosphoglycerate kinase 1 hypoxia response element (x3)、Hypoxia response element (x3) from PGK1 promoter、HRE (x3) |
| 物种来源: | H. sapiens (human) |
| 插入大小: | 79 bp |
| 基因突变: | insertion of the PGK1 HRE (x3) into promoter region |
| GenBank编号 | NM_000291 |
| 启动子: | inserted PGK1 hypoxia response elements (HRE) (x3) |
| 标签/融合蛋白: | drives expression of destabilized luciferase (2CP) for real-time HRE activity assessment (C terminal on insert) |
| 克隆方法: | 限制性内切酶 |
| 5’克隆位点: | KpnI (not destroyed) |
| 3’克隆位点: | XhoI (not destroyed) |
| 5’测序引物 | RVprimer3: CTAGCAAAATAGGCTGTCCC |
|
文献引用方法 |
|
| 材料和方法部分: | pGL4.22-PGK1-HRE::dLUC was a gift from Chi Van Dang (Addgene plasmid # 128095 ; http://n2t.net/addgene:128095 ; RRID:Addgene_128095) |
| 参考文献部分: |
Acid Suspends the Circadian Clock in Hypoxia through Inhibition of mTOR. Walton ZE, Patel CH, Brooks RC, Yu Y, Ibrahim-Hashim A, Riddle M, Porcu A, Jiang T, Ecker BL, Tameire F, Koumenis C, Weeraratna AT, Welsh DK, Gillies R, Alwine JC, Zhang L, Powell JD, Dang CV. Cell. 2018 Jun 28;174(1):72-87.e32. doi: 10.1016/j.cell.2018.05.009. Epub 2018 May 31. 10.1016/j.cell.2018.05.009 PubMed 29861175
|
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pGL4.22-PGK1-HRE::dLUC载体质粒基本信息
pGL4.22-PGK1-HRE::dLUC质粒图谱载体图谱和pGL4.22-PGK1-HRE::dLUC载体序列质粒序列多克隆位点信息
pGL4.22-PGK1-HRE::dLUC质粒载体简介
pGL4.22-PGK1-HRE::dLUC质粒序列载体序列
LOCUS pGL4.22-PGK1-HRE::dLUC 5647 bp DNA circular SYN 25-AUG-2024
DEFINITION Hypoxia response element real-time luciferase reporter.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5647)
AUTHORS Walton ZE, Patel CH, Brooks RC, Yu Y, Ibrahim-Hashim A, Riddle M,
Porcu A, Jiang T, Ecker BL, Tameire F, Koumenis C, Weeraratna AT,
Welsh DK, Gillies R, Alwine JC, Zhang L, Powell JD, Dang CV
TITLE Acid Suspends the Circadian Clock in Hypoxia through Inhibition of
mTOR.
JOURNAL Cell. 2018 Jun 28;174(1):72-87.e32. doi: 10.1016/j.cell.2018.05.009.
Epub 2018 May 31.
PUBMED 29861175
REFERENCE 2 (bases 1 to 5647)
AUTHORS .
TITLE Direct Submission
JOURNAL Exported Aug 25, 2024 from SnapGene Server 7.0.3
https://www.biofeng.com
COMMENT SGRef: number: 1; type: "Journal Article"; doi:
"10.1016/j.cell.2018.05.009"; journalName: "Cell"; date:
"2018-06-28- 28"; volume: "174"; issue: "1"; pages: "72-87.e32"
FEATURES Location/Qualifiers
source 1..5647
/mol_type="other DNA"
/organism="synthetic DNA construct"
misc_feature 1..92
/label=pause site
/note="RNA polymerase II transcriptional pause signal from
the human alpha-2 globin gene"
primer_bind 41..60
/label=RVprimer3
/note="pGL3 vector, forward primer"
regulatory 250..259
/label=Kozak sequence
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
/regulatory_class="other"
CDS 256..1905
/codon_start=1
/product="firefly luciferase"
/label=luciferase
/note="synthetic luc2 version of the luciferase gene"
/translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
CDS 1912..1959
/codon_start=1
/product="non-ORF yeast peptide conferring
ubiquitin-dependent degradation"
/label=hCL1
/note="human codon-optimized"
/translation="ACKNWFSSLSHFVIHL"
CDS 1963..2082
/codon_start=1
/product="PEST degradation sequence from mouse ornithine
decarboxylase"
/label=hPEST
/note="human codon-optimized"
/translation="SHGFPPEVEEQAAGTLPMSCAQESGMDRHPAACASARINV"
polyA_signal 2131..2252
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
primer_bind complement(2142..2161)
/label=EBV-rev
/note="SV40 polyA terminator, reverse primer"
primer_bind 2196..2215
/label=SV40pA-R
/note="SV40 polyA, reverse primer"
promoter 2446..2803
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
enhancer 2446..2637
/label=SV40 enhancer
/note="enhancer for the SV40 early promoter (Herr, 1993)"
primer_bind complement(2469..2489)
/label=pBABE 3'
/note="SV40 enhancer, reverse primer for pBABE vectors"
rep_origin 2654..2789
/label=SV40 ori
/note="SV40 origin of replication"
primer_bind 2716..2735
/label=SV40pro-F
/note="SV40 promoter/origin, forward primer"
CDS 2834..3433
/codon_start=1
/product="puromycin N-acetyltransferase"
/label=PuroR
/note="confers resistance to puromycin"
/translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
polyA_signal 3458..3506
/note="synthetic polyadenylation signal"
primer_bind complement(3662..3679)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(3833..4421)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(3913..3932)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(4621..5481)
/codon_start=1
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
polyA_signal 5586..5634
/note="synthetic polyadenylation signal"
ORIGIN
1 aacatacgct ctccatcaaa acaaaacgaa acaaaacaaa ctagcaaaat aggctgtccc
61 cagtgcaagt gcaggtgcca gaacatttct ctggcctaac tggccggtac cgagctctgt
121 cacgtcctgc acgactctag ttgtcacgtc ctgcacgact ctagttgtca cgtcctgcac
181 gacgctagct cgaggatatc aagatctggc ctcggcggcc aagcttggca atccggtact
241 gttggtaaag ccaccatgga agatgccaaa aacattaaga agggcccagc gccattctac
301 ccactcgaag acgggaccgc cggcgagcag ctgcacaaag ccatgaagcg ctacgccctg
361 gtgcccggca ccatcgcctt taccgacgca catatcgagg tggacattac ctacgccgag
421 tacttcgaga tgagcgttcg gctggcagaa gctatgaagc gctatgggct gaatacaaac
481 catcggatcg tggtgtgcag cgagaatagc ttgcagttct tcatgcccgt gttgggtgcc
541 ctgttcatcg gtgtggctgt ggccccagct aacgacatct acaacgagcg cgagctgctg
601 aacagcatgg gcatcagcca gcccaccgtc gtattcgtga gcaagaaagg gctgcaaaag
661 atcctcaacg tgcaaaagaa gctaccgatc atacaaaaga tcatcatcat ggatagcaag
721 accgactacc agggcttcca aagcatgtac accttcgtga cttcccattt gccacccggc
781 ttcaacgagt acgacttcgt gcccgagagc ttcgaccggg acaaaaccat cgccctgatc
841 atgaacagta gtggcagtac cggattgccc aagggcgtag ccctaccgca ccgcaccgct
901 tgtgtccgat tcagtcatgc ccgcgacccc atcttcggca accagatcat ccccgacacc
961 gctatcctca gcgtggtgcc atttcaccac ggcttcggca tgttcaccac gctgggctac
1021 ttgatctgcg gctttcgggt cgtgctcatg taccgcttcg aggaggagct attcttgcgc
1081 agcttgcaag actataagat tcaatctgcc ctgctggtgc ccacactatt tagcttcttc
1141 gctaagagca ctctcatcga caagtacgac ctaagcaact tgcacgagat cgccagcggc
1201 ggggcgccgc tcagcaagga ggtaggtgag gccgtggcca aacgcttcca cctaccaggc
1261 atccgccagg gctacggcct gacagaaaca accagcgcca ttctgatcac ccccgaaggg
1321 gacgacaagc ctggcgcagt aggcaaggtg gtgcccttct tcgaggctaa ggtggtggac
1381 ttggacaccg gtaagacact gggtgtgaac cagcgcggcg agctgtgcgt ccgtggcccc
1441 atgatcatga gcggctacgt taacaacccc gaggctacaa acgctctcat cgacaaggac
1501 ggctggctgc acagcggcga catcgcctac tgggacgagg acgagcactt cttcatcgtg
1561 gaccggctga agagcctgat caaatacaag ggctaccagg tagccccagc cgaactggag
1621 agcatcctgc tgcaacaccc caacatcttc gacgccgggg tcgccggcct gcccgacgac
1681 gatgccggcg agctgcccgc cgcagtcgtc gtgctggaac acggtaaaac catgaccgag
1741 aaggagatcg tggactatgt ggccagccag gttacaaccg ccaagaagct gcgcggtggt
1801 gttgtgttcg tggacgaggt gcctaaagga ctgaccggca agttggacgc ccgcaagatc
1861 cgcgagattc tcattaaggc caagaagggc ggcaagatcg ccgtgaattc tgcttgcaag
1921 aactggttca gtagcttaag ccactttgtg atccacctta acagccacgg cttccctccc
1981 gaggtggagg agcaggccgc cggcaccctg cccatgagct gcgcccagga gagcggcatg
2041 gatagacacc ctgctgcttg cgccagcgcc aggatcaacg tctaactgca gtctagagtc
2101 ggggcggccg gccgcttcga gcagacatga taagatacat tgatgagttt ggacaaacca
2161 caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat
2221 ttgtaaccat tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt
2281 ttcaggttca gggggaggtg tgggaggttt tttaaagcaa gtaaaacctc tacaaatgtg
2341 gtaaaatcga taaggatccg tttgcgtatt gggcgctctt ccgctgatct gcgcagcacc
2401 atggcctgaa ataacctctg aaagaggaac ttggttagct accttctgag gcggaaagaa
2461 ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag
2521 aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc
2581 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc
2641 cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg
2701 ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg agctattcca
2761 gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcg attcttctga
2821 cactagcgcc accatgaccg agtacaagcc taccgtgcgc ctggccactc gcgatgatgt
2881 gccccgcgcc gtccgcactc tggccgccgc tttcgccgac taccccgcta cccggcacac
2941 cgtggacccc gaccggcaca tcgagcgtgt gacagagttg caggagctgt tcctgacccg
3001 cgtcgggctg gacatcggca aggtgtgggt agccgacgac ggcgcggccg tggccgtgtg
3061 gactaccccc gagagcgttg aggccggcgc cgtgttcgcc gagatcggcc cccgaatggc
3121 cgagctgagc ggcagccgcc tggccgccca gcagcaaatg gagggcctgc ttgcccccca
3181 tcgtcccaag gagcctgcct ggtttctggc cactgtagga gtgagccccg accaccaggg
3241 caagggcttg ggcagcgccg tcgtgttgcc cggcgtagag gccgccgaac gcgccggtgt
3301 gcccgccttt ctcgaaacaa gcgcaccaag aaaccttcca ttctacgagc gcctgggctt
3361 caccgtgacc gccgatgtcg aggtgcccga gggacctagg acctggtgta tgacacgaaa
3421 acctggcgcc taatgatcta gaaccggtca tggccgcaat aaaatatctt tattttcatt
3481 acatctgtgt gttggttttt tgtgtgttcg aactagatgc tgtcgaccga tgcccttgag
3541 agccttcaac ccagtcagct ccttccggtg ggcgcggggc atgactatcg tcgccgcact
3601 tatgactgtc ttctttatca tgcaactcgt aggacaggtg ccggcagcgc tcttccgctt
3661 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
3721 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
3781 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
3841 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
3901 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
3961 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
4021 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
4081 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
4141 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
4201 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
4261 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
4321 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
4381 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
4441 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
4501 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
4561 aaagtatata tgagtaaact tggtctgaca gcggccgcaa atgctaaacc actgcagtgg
4621 ttaccagtgc ttgatcagtg aggcaccgat ctcagcgatc tgcctatttc gttcgtccat
4681 agtggcctga ctccccgtcg tgtagatcac tacgattcgt gagggcttac catcaggccc
4741 cagcgcagca atgatgccgc gagagccgcg ttcaccggcc cccgatttgt cagcaatgaa
4801 ccagccagca gggagggccg agcgaagaag tggtcctgct actttgtccg cctccatcca
4861 gtctatgagc tgctgtcgtg atgctagagt aagaagttcg ccagtgagta gtttccgaag
4921 agttgtggcc attgctactg gcatcgtggt atcacgctcg tcgttcggta tggcttcgtt
4981 caactctggt tcccagcggt caagccgggt cacatgatca cccatattat gaagaaatgc
5041 agtcagctcc ttagggcctc cgatcgttgt cagaagtaag ttggccgcgg tgttgtcgct
5101 catggtaatg gcagcactac acaattctct taccgtcatg ccatccgtaa gatgcttttc
5161 cgtgaccggc gagtactcaa ccaagtcgtt ttgtgagtag tgtatacggc gaccaagctg
5221 ctcttgcccg gcgtctatac gggacaacac cgcgccacat agcagtactt tgaaagtgct
5281 catcatcggg aatcgttctt cggggcggaa agactcaagg atcttgccgc tattgagatc
5341 cagttcgata tagcccactc ttgcacccag ttgatcttca gcatctttta ctttcaccag
5401 cgtttcgggg tgtgcaaaaa caggcaagca aaatgccgca aagaagggaa tgagtgcgac
5461 acgaaaatgt tggatgctca tactcgtcct ttttcaatat tattgaagca tttatcaggg
5521 ttactagtac gtctctcaag gataagtaag taatattaag gtacgggagg tattggacag
5581 gccgcaataa aatatcttta ttttcattac atctgtgtgt tggttttttg tgtgaatcga
5641 tagtact
//
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