Abstract:
:To investigate the feasibility of conducting a genomic-scale protein labeling and localization study in Escherichia coli, a representative subset of 23 coding DNA sequences (CDSs) was selected for chromosomal tagging with one or more fluorescent protein genes (EGFP, EYFP, mRFP1, DsRed2). We used lambda-Red recombination to precisely and efficiently position PCR-generated DNA targeting cassettes containing a fluorescent protein gene and an antibiotic resistance marker, at the C-termini of the CDSs of interest, creating in-frame fusions under the control of their native promoters. We incorporated cre/loxP and flpe/frt technology to enable multiple rounds of chromosomal tagging events to be performed sequentially with minimal disruption to the target locus, thus allowing sets of proteins to be co-localized within the cell. The visualization of labeled proteins in live E. coli cells using fluorescence microscopy revealed a striking variety of distributions including: membrane and nucleoid association, polar foci and diffuse cytoplasmic localization. Fifty of the fifty-two independent targeting experiments performed were successful, and 21 of the 23 selected CDSs could be fluorescently visualized. Our results show that E. coli has an organized and dynamic proteome, and demonstrate that this approach is applicable for tagging and (co-) localizing CDSs on a genome-wide scale.
journal_name
Nucleic Acids Resjournal_title
Nucleic acids researchauthors
Watt RM,Wang J,Leong M,Kung HF,Cheah KS,Liu D,Danchin A,Huang JDdoi
10.1093/nar/gkl1158subject
Has Abstractpub_date
2007-01-01 00:00:00pages
e37issue
6eissn
0305-1048issn
1362-4962pii
gkl1158journal_volume
35pub_type
杂志文章abstract::Chlorella virus PBCV-1 DNA ligase seals nicked DNA substrates consisting of a 5'-phosphate-terminated strand and a 3'-hydroxyl-terminated strand annealed to a bridging DNA template strand. The enzyme discriminates at the DNA binding step between substrates containing a 5'-phosphate versus a 5'-hydroxyl at the nick. Mu...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/26.2.525
更新日期:1998-01-15 00:00:00
abstract::We have studied a novel class of DNA sequences that cause DNA polymerases to pause. These sequences have the central consensus Py-G-C and are not necessarily adjacent to hairpins in the DNA template. Since most consensus sequences do not cause pauses under standard conditions, additional template features must exist t...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/24.14.2774
更新日期:1996-07-15 00:00:00
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 e...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gni055
更新日期:2005-03-22 00:00:00
abstract::Genome-wide association (GWA) studies to map genes for complex traits are powerful yet costly. DNA-pooling strategies have the potential to dramatically reduce the cost of GWA studies. Pooling using Affymetrix arrays has been proposed and used but the efficiency of these arrays has not been quantified. We compared and...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gkm1060
更新日期:2008-04-01 00:00:00
abstract::MDg3 is a family of mobile dispersed genetic elements represented by 15 copies in the haploid genome of D. melanogaster and flanked, like other similar elements, by the regions of homology. In the present work, these regions of mdg3 have been sequenced. The existence of perfect direct repeats 268 base pairs long has b...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/8.15.3263
更新日期:1980-08-11 00:00:00
abstract::A method is presented for the purification of mRNAs and the identification of structural gene sequences in recombinant DNA molecules. RNA is hybridized to double-stranded linear DNA such that R-loops are formed between most DNAs and their complementary RNA sequences. These R-loops are purified from unhybridized RNAs b...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/6.7.2483
更新日期:1979-06-11 00:00:00
abstract::IMP (Integrative Multi-species Prediction), originally released in 2012, is an interactive web server that enables molecular biologists to interpret experimental results and to generate hypotheses in the context of a large cross-organism compendium of functional predictions and networks. The system provides biologists...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gkv486
更新日期:2015-07-01 00:00:00
abstract::Unwanted side effects of drugs are a burden on patients and a severe impediment in the development of new drugs. At the same time, adverse drug reactions (ADRs) recorded during clinical trials are an important source of human phenotypic data. It is therefore essential to combine data on drugs, targets and side effects...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gkv1075
更新日期:2016-01-04 00:00:00
abstract::TORQUE is a tool for cross-species querying of protein-protein interaction networks. It aims to answer the following question: given a set of proteins constituting a known complex or a pathway in one species, can a similar complex or pathway be found in the protein network of another species? To this end, Torque seeks...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gkp474
更新日期:2009-07-01 00:00:00
abstract::The rpoA gene, encoding the alpha subunit of RNA polymerase, was cloned from Streptomyces coelicolor A3(2). It is preceded by rpsK and followed by rplQ, encoding ribosomal proteins S11 and L17, respectively, similar to the gene order in Bacillus subtilis. The rpoA gene specifies a protein of 339 amino acids with deduc...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/24.22.4565
更新日期:1996-11-15 00:00:00
abstract::The PD-(D/E)XK type II restriction endonuclease ThaI cuts the target sequence CG/CG with blunt ends. Here, we report the 1.3 Å resolution structure of the enzyme in complex with substrate DNA and a sodium or calcium ion taking the place of a catalytic magnesium ion. The structure identifies Glu54, Asp82 and Lys93 as t...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gkq834
更新日期:2011-01-01 00:00:00
abstract::We have sequenced mouse tRNA genes from two recombinant lambda phage. An 1800 bp sequence from one phage contains 3 tRNA genes, potentially encoding tRNAAsp, tRNAGly, and tRNAGlu, separated by spacer sequences of 587 bp and 436 bp, respectively. The mouse tRNA gene cluster is homologous to a rat sequence (Sekiya et al...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/11.24.8761
更新日期:1983-12-20 00:00:00
abstract::The signal recognition particle database (SRPDB) is located at the University of Texas Health Science Center at Tyler and includes tabulations of SRP RNA, SRP protein and SRP receptor sequences. The sequences are annotated with links to the primary databases. They are ordered alphabetically or phylogenetically and are...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/27.1.169
更新日期:1999-01-01 00:00:00
abstract::We have exploited properties of photosensitizers to study an aspect of the packing of chromatin in the cell nucleus. The fluorescent photosensitizers mesotetra(3-hydroxyphenyl) porphyrin and Photofrin II were both localized in the nuclear membrane and other membrane structures, but could not be found inside the nuclei...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/20.24.6687
更新日期:1992-12-25 00:00:00
abstract::Chewie Nomenclature Server (chewie-NS, https://chewbbaca.online/) allows users to share genome-based gene-by-gene typing schemas and to maintain a common nomenclature, simplifying the comparison of results. The combination between local analyses and a public repository of allelic data strikes a balance between potenti...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gkaa889
更新日期:2021-01-08 00:00:00
abstract::We recently developed a new type of pyrrole (Py)-imidazole (Im) polyamide-tetrahydrocyclopropabenzindolone (CBI) conjugate with an indole linker as a stable sequence-specific alkylating agent. In this study, we investigated the gene silencing activities of polyamides A, B and C, which selectively alkylate specific seq...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gkl005
更新日期:2006-02-25 00:00:00
abstract::The role of the cyclic-AMP (cAMP) responsive element (CRE) in eukaryotic gene transcription was investigated in several cell lines transfected by constructs containing the chloramphenicol acetyltransferase (CAT) gene linked to the three different promoters, simian virus (SV) 40, human c-Ha-ras-1, or chicken beta-actin...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/17.4.1521
更新日期:1989-02-25 00:00:00
abstract::We have isolated from a single human liver cDNA library two clones which are highly homologous (78% over the coding region) to the major phenobarbital-inducible P450 from rat (P450IIB1). This is the first direct demonstration of the presence of the P450IIB gene subfamily in humans. This subfamily is much less extensiv...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/16.13.5783
更新日期:1988-07-11 00:00:00
abstract::Primer design is a fundamental technique that is widely used for polymerase chain reaction (PCR). Although many methods have been proposed for primer design, they require a great deal of manual effort to generate feasible and valid primers, including homology tests on off-target sequences using BLAST-like tools. That ...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gkv632
更新日期:2015-11-16 00:00:00
abstract::Recombinant plasmids which contain EcoRI fragments of tobacco chloroplast DNA carrying tRNA genes were constructed. Plasmids pTC211 and pTC293 contain the base sequences for tRNAAsn in their 1.4 and 1.1 Md EcoRI fragments, respectively. These two tRNA sequences are identical and are; 5'-TCCTCAGTAGCTCAGTGGTAGAGCGGTCGGC...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/9.21.5601
更新日期:1981-11-11 00:00:00
abstract::We have measured the amount of 5S-ribosomal DNA in the genomes of Xenopus laevis, Triturus cristatus carnifex and Ambystoma mexicanum, three species of Amphibians which have widely different C-values. Our best estimate is that these organisms have about 24,000, 32,000 and 61,000 5S-genes per haploid genome respectivel...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/11.8.2381
更新日期:1983-04-25 00:00:00
abstract::The Pk-rec gene, encoding a RecA/RAD51 homologue from the hyperthermophilic archaeon Pyrococcussp. KOD1, was expressed in Escherichia coli. The recombinant Pk-REC was purified to homogeneity and was shown to be in a dimeric form. A striking property of the purified recombinant Pk-REC was the unusual DNase activity on ...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/25.4.719
更新日期:1997-02-15 00:00:00
abstract::Mitotic chromosome structure is pivotal to cell division but difficult to observe in fine detail using conventional methods. DNA catenation has been implicated in both sister chromatid cohesion and chromosome condensation, but has never been observed directly. We have used a lab-on-a-chip microfluidic device and fluor...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gks931
更新日期:2012-12-01 00:00:00
abstract::The underlying mechanism of transcriptional co-repressor ETO2 during early erythropoiesis and hemoglobin switching is unclear. We find that absence of ETO2 in mice interferes with down-regulation of PU.1 and GATA2 in the fetal liver, impeding a key step required for commitment to erythroid maturation. In human β-globi...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gkaa736
更新日期:2020-10-09 00:00:00
abstract::We created a single cell sorting system to screen for enzyme activity in Escherichia coli producing 3,4 dihydroxy benzoate (34DHB). To do so, we engineered a transcription factor regulon controlling the expression of green fluorescent protein (GFP) for induction by 34DHB. An autoregulated transcription factor, pcaU, w...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gku444
更新日期:2014-07-01 00:00:00
abstract::The EMBL Data Library was the first internationally supported central resource for nucleic acid sequence data. Working in close collaboration with its American counterpart, GenBank (1), the library prepares and makes available to the scientific community a comprehensive collection of the published nucleic acid sequenc...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/14.1.5
更新日期:1986-01-10 00:00:00
abstract::The sequences coding for DNA[cytosine-N4]methyltransferases MvaI (from Micrococcus varians RFL19) and Cfr9I (from Citrobacter freundii RFL9) have been determined. The predicted methylases are proteins of 454 and 300 amino acids, respectively. Primary structure comparison of M.Cfr9I and another m4C-forming methylase, M...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/17.23.9823
更新日期:1989-12-11 00:00:00
abstract::The 279-bp major breakpoint region (mbr) within the 3'-untranslated region (3'-UTR) of the BCL2 gene is a binding site of special AT-rich sequence binding protein 1 (SATB1) that is well known to participate in the long-range regulation of gene transcription. Our previous studies have revealed that the mbr could regula...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gkr023
更新日期:2011-06-01 00:00:00
abstract::Exome sequencing strategy is promising for finding novel mutations of human monogenic disorders. However, pinpointing the casual mutation in a small number of samples is still a big challenge. Here, we propose a three-level filtration and prioritization framework to identify the casual mutation(s) in exome sequencing ...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gkr1257
更新日期:2012-04-01 00:00:00
abstract::Most RNA molecules collapse rapidly and reach the native state through a pathway that contains numerous traps and unproductive intermediates. The D135 group II intron ribozyme is unusual in that it can fold slowly and directly to the native state, despite its large size and structural complexity. Here we use hydroxyl ...
journal_title:Nucleic acids research
pub_type: 杂志文章
doi:10.1093/nar/gki973
更新日期:2005-11-27 00:00:00