Abstract:
:In a gene targeting experiment, the generation of a targeting construct often requires complex DNA manipulations. We developed a set of cassettes and plasmids useful for creating targeting vectors to modify the mammalian genome. A positive selection marker cassette (PGK/EM7p-npt), which included dual prokaryotic and eukaryotic promoters to permit consecutive selection for recombination in Escherichia coli and then in mouse embryonic stem cells, was flanked by two FRT-loxP sequences. The PGK/EM7p-npt cassette was placed between the minimum regions of a Tn7 transposable element for insertion into another DNA by means of Tn7 transposase in vitro. We also constructed a plasmid having a loxP-Zeo-loxP cassette between the modified Tn5 outer elements. These cassettes can be integrated randomly into a given genomic DNA through the in vitro transposition reaction, thus producing a collection of genomic segments flanked by loxP sites (floxed) at various positions without the use of restriction enzymes and DNA ligase. We confirmed that this system remarkably reduced the time and labor for the construction of complex gene targeting vectors.
journal_name
Nucleic Acids Resjournal_title
Nucleic acids researchauthors
Aoyama M,Agari K,Sun-Wada GH,Futai M,Wada Ydoi
10.1093/nar/gni055keywords:
subject
Has Abstractpub_date
2005-03-22 00:00:00pages
e52issue
5eissn
0305-1048issn
1362-4962pii
33/5/e52journal_volume
33pub_type
杂志文章abstract::An investigation of the mobility of short duplexes of DNA -octamers and decamers-on polyacrylamide gels is presented, showing that molecules containing less than one helical turn exhibit sequence dependent mobilities. Analysis of chains with different sequences indicates that any arrangement of two or more adjacent A....
journal_title:Nucleic acids research
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abstract::This report documents the error rate in a commercially distributed subset of the IMAGE Consortium mouse cDNA clone collection. After isolation of plasmid DNA from 1189 bacterial stock cultures, only 62. 2% were uncontaminated and contained cDNA inserts that had significant sequence identity to published data for the o...
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pub_type: 杂志文章
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更新日期:1983-03-11 00:00:00
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journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gki474
更新日期:2005-07-01 00:00:00
abstract::The human transcription factor EF-1A binds to the purine-rich E1A core enhancer sequence in the adenovirus E1A and E4 and polyomavirus enhancer regions. The consensus binding site for EF-1A resembles that of members of the ets domain protein family. EF-1A activation of transcription requires a dimeric binding site. An...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/20.24.6555
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journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gkm661
更新日期:2007-01-01 00:00:00
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journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/17.20.8221
更新日期:1989-10-25 00:00:00
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journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/11.20.7169
更新日期:1983-10-25 00:00:00
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journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gkp051
更新日期:2009-04-01 00:00:00
abstract::In Eukaryotes, DNA is wound around the histone octamer forming the basic chromatin unit, the nucleosome. Atomic structures have been obtained from crystallography and single particle cryo-electron microscopy (cryoEM) of identical engineered particles. But native nucleosomes are dynamical entities with diverse DNA sequ...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gky670
更新日期:2018-09-28 00:00:00
abstract::The use of the DNA duplex as a supramolecular scaffold is an established approach for the assembly of chromophore aggregates. In the absence of detailed structural insight, the characterization of thus assembled oligochromophores is, today, largely based on solution-phase spectroscopy. Here, we describe the crystal st...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gkw644
更新日期:2016-09-06 00:00:00
abstract::'Linker scanning' has been used as an efficient method for systematically surveying a segment of DNA for functional elements by mutagenesis. A three-step PCR method was developed to simplify this process. In this method, a set of 'mutation primers' was made with 6 to 8 base substitutions in the center of the primers. ...
journal_title:Nucleic acids research
pub_type: 杂志文章
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更新日期:1993-08-11 00:00:00
abstract::We have used in vitro site-directed mutagenesis with synthetic DNA oligonucleotides to introduce single nucleotide mutations in yeast mtDNA. In addition to the expected DNA alterations we also recovered with high frequency mutants with large deletions and insertions which arose through interaction with the synthetic D...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/11.24.8595
更新日期:1983-12-20 00:00:00
abstract::IGF2 mRNA-binding protein 1 (IMP1) is a key regulator of messenger RNA (mRNA) metabolism and transport in organismal development and, in cancer, its mis-regulation is an important component of tumour metastasis. IMP1 function relies on the recognition of a diverse set of mRNA targets that is mediated by the combinator...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gkz136
更新日期:2019-05-07 00:00:00
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journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/24.4.754
更新日期:1996-02-15 00:00:00
abstract::The plastid psbA promoter of tobacco was used with the aim to construct plastid specific marker genes. Upon transfer to the tobacco nuclear genome the plastid promoter fragment appeared to specify a messenger RNA. Placed 5' to the bar or nptII coding sequences the level of expression is sufficient to obtain a selectab...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/17.1.19
更新日期:1989-01-11 00:00:00
abstract::Hepatitis B virus (HBV) infection remains a major health problem worldwide. The role played by microRNAs (miRNAs) in HBV replication and pathogenesis is being increasingly recognized. In this study, we found that miR-15b, an important miRNA during HBV infection and hepatocellular carcinoma development, directly binds ...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gku260
更新日期:2014-06-01 00:00:00
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journal_title:Nucleic acids research
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journal_title:Nucleic acids research
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journal_title:Nucleic acids research
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journal_title:Nucleic acids research
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更新日期:2006-01-01 00:00:00
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journal_title:Nucleic acids research
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journal_title:Nucleic acids research
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journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/11.3.629
更新日期:1983-02-11 00:00:00
abstract::Selection for amino acid affinity by elution of RNAs from tryptophan-Sepharose using free L-tryptophan evokes one sequence predominantly (K(D) = 12 microM), a symmetrical internal loop of 3 nt per side. Though we have also isolated larger sequences with affinity for tryptophan, successively squeezed selection in rando...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gki861
更新日期:2005-09-25 00:00:00
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journal_title:Nucleic acids research
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journal_title:Nucleic acids research
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journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gkq943
更新日期:2011-01-01 00:00:00
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journal_title:Nucleic acids research
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doi:10.1093/nar/17.3.1051
更新日期:1989-02-11 00:00:00