No data available that match "Blotting, Southern"



*  Southern Blotting | Thermo Fisher Scientific

Southern blot analysis reveals information about DNA identity, size, and abundance. It is a classic technique that involves ... Southern blot analysis reveals information about DNA identity, size, and abundance. It is a classic technique that involves ... Obtaining complete fragmentation of your DNA at the intended restriction enzyme sites is a critical step in Southern blot ... unambiguous Southern blot data. Each restriction enzyme has been validated for use with one of our universal buffers (L, M, H, ...
thermofisher.com/de/de/home/life-science/dna-rna-purification-analysis/nucleic-acid-gel-electrophoresis/southern-blotting.html

*  Question about Southern blotting - General Lab Techniques

Question about Southern blotting - (Jul/30/2012 ). Hii. Where can I order the paper towels for southern blotting ? ...
protocol-online.org/biology-forums-2/posts/26367.html

*  Norther Blotting. Southern Blotting. Western Blotting. | Learning Material | Noodle

Southern Blotting. Western Blotting.. See what people are saying about Norther Blotting. Southern Blotting. Western Blotting.. ... Get an in-depth review and ask questions about Norther Blotting. ... Southern Blotting. Western Blotting.. Want more info about ... Southern Blotting. Western Blotting.? Get free advice from education experts and Noodle community members. ... Norther Blotting. Southern Blotting. Western Blotting.. http://www.dnaPot.com Follow us on twitter: http://www.twitter.com/#!/ ...
https://noodle.com/learn/details/17540/norther-blotting-southern-blotting-western-blotting

*  Northern & Southern Blot

Make probes for northern blotting and southern blotting using our labeled nucleotides. Ensure reliable results in your DNA and ... RNA blotting experiments through our wide selection of transfer membranes, enzyme conjugates and chemilumescent detection ...
perkinelmer.co.uk/category/northern-southern-blot

*  Best 20+ Southern blot ideas on Pinterest | Makeup vanity set, Vanities and Makeup vanities ideas

Find and save ideas about Southern blot on Pinterest. , See more ideas about Makeup vanity set, Vanities and Makeup vanities ... Southern blotting (named after Edwin M. Southern) is one technique used to transfer the denatured, single-stranded fragments ... Southern, Northern, and Western blot mnemonic medicowesome: "medicowesome: " Hello everyone! The mnemonic to remember blotting ... Southern BlotLearn ScienceMedical LaboratoryMolecular BiologySchool HacksBiotechnologyGeneticsPoolsWorkshop. ...
https://pinterest.co.uk/explore/southern-blot/

*  What is Southern Blotting? How it is used to identify DNA Sequences?

Southern Blotting is an elegant technique that can be used to identify the DNA sequences on restriction fragments separated by ... What is Southern Blotting? How it is used to identify Specific DNA Sequences?. March 29, 2016. Vinod Thakur Leave a comment ... Southern Blotting is an elegant technique that can be used to identify the locations of genes and other DNA sequences on ... Applications of Southern Blotting:. *An invaluable method for gene analysis and confirming DNA cloning results. ...
https://sciencesamhita.com/what-is-southern-blotting-its-procedure-applications/

*  Gene replacement strategy and Southern blot analysis of | Open-i

Gene replacement strategy and Southern blot analysis of progeny from heterozygous crosses. (a) The wild-type P-selectin allele ... A probe flanking the 3′-end of the vector was used for Southern blot analysis of ES cell clones and tail biopsies of the ... A probe flanking the 3′-end of the vector was used for Southern blot analysis of ES cell clones and tail biopsies of the ... b) Representative Southern blot analysis of tail biopsies of progeny from heterozygous crosses. Genomic DNA was digested with ...
https://openi.nlm.nih.gov/detailedresult.php?img=PMC2132959_JCB9805113.f1b&req=4

*  Changes of telomere length with aging - Takubo - 2010 - Geriatrics & Gerontology International - Wiley Online Library

Prior studies of telomere length in breast cancers using Southern blotting or slot-blot analysis have shown telomere shortening ... 52 These disagreements might be partly explained by the limitations of Southern blotting and slot-blot analysis, which are ... In previous studies by Kang et al.40 and our group,41 the Southern blotting technique was used, and Meeker et al.42 used Q-FISH ... Southern blotting. Subjects, tissues and DNA samples. When we used tissues from autopsied subjects, all the cadavers examined ...
onlinelibrary.wiley.com/doi/10.1111/j.1447-0594.2010.00605.x/full

*  Leukemia - MLL-AF9 and FLT3 cooperation in acute myelogenous leukemia: development of a model for rapid therapeutic assessment

Southern blots were probed with the EGFP sequence to determine the clonality of the transplanted diseases. Shown are Southern ... Southern blot confirmed oligoclonality of both diseases (. Figure 3). Thus, addition of FLT3-ITD does not alter the leukemic ... DNA extraction and Southern blotting. Genomic DNA was extracted from spleens using the Gentra Systems kit (Gentra Systems, ... Western blots were performed using the 4G10 mouse -phosphotyrosine, goat -hFLT3 (R&D Systems) or mouse -pan- actin (Chemicon, ...
nature.com/leu/journal/v22/n1/full/2404951a.html?foxtrotcallback=true&error=cookies_not_supported&code=08e5025f-44d8-4878-8cab-bd7d6d08450b

*  The Role of Stn1p in Saccharomyces cerevisiae Telomere Capping Can Be Separated From Its Interaction With Cdc13p | Genetics

For both Southern blots, the genomic DNA was digested with XhoI and the membrane was probed with [32P]d(GT/CA). (A) Southern ... A) Southern blot of stn1-281t, stn1-186t, and stn1-186t rad52-Δ strains (hC671, hC672, hC1110). Genomic DNA was prepared from ... A) Southern blot showing telomere lengths in diploids created by mating stn1-186t with est2-Δ (three dCN325 diploid strains ... Southern blots:. Cell cultures were grown to saturation at 30°, and genomic DNA was prepared as previously described (Lundblad ...
genetics.org/content/177/3/1459

*  Trans-Kingdom Transposition of the Maize Dissociation Element | Genetics

Southern blot hybridization:. Genomic DNA from pooled zebrafish embryos was phenol extracted and digested using the EcoRI ... Southern blot hybridization with EGFP-specific probe revealed predominantly multiple insertions in the genome of individual F1 ... was outcrossed to a wild-type fish and DNA from 12 pooled randomly selected EGFP-positive F2 embryos was used for Southern blot ... by capillary blotting (Sambrook et al. 1989), and crosslinked by UV irradiation. The DNA probe for EGFP was labeled with ...
genetics.org/content/174/3/1095

*  A transgenic mouse model demonstrating the oncogenic role of mutations in the polycomb-group gene EZH2 in lymphomagenesis |...

Southern blots for determination of clonality of the V(D)J rearrangement were performed by digesting the DNA with EcoRI ( ... F) Southern blot analysis for clonality of IgH gene rearrangements for Eµ-Myc lymphomas (upper) and Eµ-Myc/EZH2Y641F lymphomas ... Southern blot. Genomic DNA was extracted from cut mouse tail tips by proteinase K digestion followed by a phenol-chloroform ... B) Southern blot using a probe specific for the human growth hormone (hGH) sequence in the transgene performed on 3 transgenic ...
bloodjournal.org/content/123/25/3914?sso-checked=true

*  Nucleic Acid Gel Electrophoresis | Thermo Fisher Scientific

DNA and RNA gel electrophoresis and blotting tools and reagents, featuring the E-Gel® Electrophoresis System. ... Southern blotting. Find products to reveal information about DNA identity, size, and abundance. ... We've developed an expansive collection of tools and reagents for DNA and RNA gel electrophoresis and blotting. You'll find all ... Northern blotting. Find products to reveal information about RNA identity, size, and abundance. ...
thermofisher.com/ca/en/home/life-science/dna-rna-purification-analysis/nucleic-acid-gel-electrophoresis.html

*  Plus it

DNA Isolation and Southern Blotting.. High molecular weight DNA was isolated by phenol/chloroform extraction. Each sample (20 ... Hybridization was performed for 20 h at 68°C (38) . Blots were analyzed with a Fuji BAS 2000 phosphoimager (Raytest, ... To determine whether GLI gene amplification may underlie the elevated expression of GLI mRNA in some samples, Southern ... and samples of 15 normal mesenchymal tissues were examined for GLI gene amplification and expression by Southern hybridization ...
cancerres.aacrjournals.org/content/59/8/1890

*  Gentamicin resistance genes in environmental bacteria: prevalence and transfer - Heuer - 2002 - FEMS Microbiology Ecology -...

Dot and Southern blotting was performed by standard protocols [25]. Hybridisations of Southern- or dot-blotted plasmid DNA or ... 2.9Dot-blot and Southern blot analysis. Genomic DNA extracted from Gm-resistant isolates or transconjugants was analysed by dot ... Hybridisation of Southern-blotted PCR products obtained with the aac(6′)-II/Ib primer system from total community DNA. First ... Southern blots of the PCR products were always probed with the PCR-derived probes for the respective gene cluster to increase ...
onlinelibrary.wiley.com/doi/10.1111/j.1574-6941.2002.tb01019.x/full

*  Suppression of Ku70/80 or Lig4 leads to decreased stable transformation and enhanced homologous recombination in rice -...

Red bar represents DNA probe for Southern blot analysis. (b) Southern blot analysis using a codA probe (shown by the red bar in ... Southern blot analysis with a CodA probe showed that almost all transformants contained a single copy or two copies of the T- ... Southern blot analysis was performed according to a standard protocol. Specific DNA probes for the codA gene were synthesized ... Genomic DNA extraction and Southern blot analysis. Genomic DNA was extracted from rice calli transformed with the LU-UC ...
onlinelibrary.wiley.com/doi/10.1111/j.1469-8137.2012.04350.x/full?globalMessage=0&systemMessage=Wiley Online Library will be unavailable on Saturday 7th Oct from 03.00 EDT / 08:00 BST / 12:30 IST / 15.00 SGT to 08.00 EDT / 13.00 BST / 17:30 IST / 20.00 SGT and Sunday 8th Oct from 03.00 EDT / 08:00 BST / 12:30 IST / 15.00 SGT to 06.00 EDT / 11.00 BST / 15:30 IST / 18.00 SGT for essential maintenance. Apologies for the inconvenience caused

*  Plus it

Southern blot analysis. Mouse genomic DNA was digested withEcoRI or BamHI, electrophoresed in a 0.7% agarose gel (7 μg/lane), ... Southern blot analysis with mGIF probe. To assess the possible presence of other genes homolgous to mGIF in the mouse genome, ... Genomic DNA Blot Analysis with mGIF probe. Southern hybridization was performed on mouse genomic DNA followed by washing at low ... Northern blot analysis. To study the distribution of mGIF, a mouse multiple tissue northern (MTN) blot membrane (Clontech, Palo ...
jneurosci.org/content/17/22/8657

*  Kallikrein Gene Delivery Inhibits Vascular Smooth Muscle Cell Growth and Neointima Formation in the Rat Artery After Balloon...

Gene-Specific Primers and Probes for Rat Kallikrein-Kinin Systems and Human Tissue Kallikrein in RT-PCR Southern Blot Analysis ... Reverse Transcription-Polymerase Chain Reaction Southern Blot Analysis. Total RNA was extracted with Trizol reagent according ... Southern blot analyses were used to determine the abundance of B2 receptors, rat tissue kallikrein, high- /and low-molecular- ... primers used for RT-PCR and specific internal oligonucleotide probes for Southern blot analyses. Signals were detected by ...
hyper.ahajournals.org/content/34/2/164

*  Study of viral integration of HPV-16 in young patients with LSIL | Journal of Clinical Pathology

Southern blot hybridisation. Southern blot hybridisation was performed on all of the 92 samples to confirm the PCR data for the ... In phase 2, Southern blot hybridisation was performed on the E6 gene PCR product as a control. Thirteen of the 92 cases were ... The use of Southern blot analysis has several advantages: it allows confirmation of the type of HPV identified, which was type ... Figure 2 shows the E2 PCR products of some of the 13 samples that were positive by both PCR and Southern blot for the E6 gene, ...
jcp.bmj.com/content/56/7/532

*  Methods - Biology Online

Plasmid isolation and Southern blotting. Extrachromosomal DNA elements were extracted from P. acidilactici UVA1, UL5 and the ... Plasmid preparation, blotting and hybridization were performed twice.. RNA isolation and reverse transcription. The RNA was ... DIG-labelling of the pedA-probe (P1-P2 PCR product on P. acidilactici UVA1), blotting on nylon membrane, hybridisation (at 42°C ...
biology-online.org/articles/detection_pediocin_gene_peda/methods.html

*  Plus it

DNA was blotted, and a labeled HNF3α probe was hybridized to the Southern filter using the protocol described previously (10) ... Southern Blot Analysis and Interphase FISH Assay.. Ten μg of high molecular weight DNA from paired normal-tumor tissues were ... Southern blot analysis was performed on three pairs of esophageal normal-tumor samples using a HNF3α-specific probe, and gene ... B, gene amplification of HNF3α is verified by Southern blot analysis using HNF3α gene-specific probe. ...
cancerres.aacrjournals.org/content/62/18/5273

*  Alteration of DNA methylation in gastrointestinal carcinogenesis - Fang - 2001 - Journal of Gastroenterology and Hepatology -...

Digestion by Hpa II/Msp I and the use of Southern blotting. Until 5 years ago, Southern blotting is the most frequently used ... the methylated status of c-myc and c-Ha-ras oncogenes in human gastric cancer with the use of Southern blotting was analyzed. ...
onlinelibrary.wiley.com/doi/10.1046/j.1440-1746.2001.02554.x/full

*  Endothelial Cells of the Human Microvasculature Express Epidermal Fatty Acid-Binding Protein | Circulation Research

Southern Blotting, Cloning, and Sequencing of PCR Products. The amplified cDNA fragments were separated in a 1.5% agarose gel ... Southern blotting using the radiolabeled PA-FABP full-length cDNA as a probe further confirmed the expression of E-FABP in ... Southern blotting and sequencing of the cloned RT-PCR products demonstrate that endothelial E-FABP is identical to keratinocyte ...
circres.ahajournals.org/content/81/3/297

*  Cooperativity between DNA Methyltransferases in the Maintenance Methylation of Repetitive Elements

Southern blot analysis of DNA from cell lines. Ten micrograms of DNA from cell lines was separately digested with either 50 U ... using either Southern blot analysis or Ms-SNuPE analysis (Fig. 1B and C). Results confirmed the decreases apparent from the AP- ... A-repeat clones were sequenced by automated DNA sequencer at the University of Southern California (USC)/Norris Comprehensive ... agarose gel and Southern transferred to Zeta-Probe (Bio-Rad) membranes overnight. Cloned DNA fragments that had previously been ...
pubmedcentralcanada.ca/pmcc/articles/PMC139739/?lang=en-ca

*  Orphanet: Simple search

Mitochondrial southern blot for detection of multiple deletions, dublications and depletions The Cyprus Institute of Neurology ... Molecular screening for Mitochondrial Diseases (mtDNA deletion screen in blood, muscle & liver tissue by southern blot) ...
orpha.net/consor/cgi-bin/ClinicalLabs_Search_Simple.php?lng=EN&LnkId=19262&Typ=Pat

No data available that match "Blotting, Southern"



(1/10141) Isolation of human transcripts expressed in hamster cells from YACs by cDNA representational difference analysis.

Gene isolation methods used during positional cloning rely on physical contigs consisting of bacterial artificial chromosomes, P1, or cosmid clones. However, in most instances, the initial framework for physical mapping consists of contigs of yeast artificial chromosome (YACs), large vectors that are suboptimal substrates for gene isolation. Here we report a strategy to identify gene sequences contained within a YAC by using cDNA representational difference analysis (RDA) to directly isolate transcripts expressed from the YAC in mammalian cells. The RDA tester cDNAs were generated from a previously reported hamster cell line derived by stable transfer of a 590-kb YAC (911D5) that expressed NPC1, the human gene responsible for Niemann-Pick type C (NP-C). The driver cDNAs were generated from a control hamster cell line that did not contain the YAC that expressed NPC1. Among the gene fragments obtained by RDA, NPC1 was the most abundant product. In addition, two non-NPC1 fragments were isolated that were mapped to and expressed from 911D5. One of these RDA gene fragments (7-R) spans more than one exon and has 98% sequence identity with a human cDNA clone reported previously as an expressed sequence tag (EST), but not mapped to a chromosomal region. The other fragment (2-R) that had no significant sequence similarities with known mammalian genes or ESTs, was further localized to the region of overlap between YACs 911D5 and 844E3. The latter YAC is part of a contig across the NP-C candidate region, but does not contain NPC1. This two-part approach in which stable YAC transfer is followed by cDNA RDA should be a useful adjunct strategy to expedite the cloning of human genes when a YAC contig is available across a candidate interval.  (+info)

(2/10141) Human papillomavirus DNA in adenosquamous carcinoma of the lung.

AIM: To investigate the presence of human papillomavirus (HPV) DNA in adenosquamous carcinoma of the lung--which is relatively common in Okinawa but not in mainland Japan--and examine its histological features. METHODS: Of 207 cases where primary lung cancers were surgically removed between January 1995 and June 1997 in Okinawa, 23 were adenosquamous carcinoma. HPV was detected by non-isotopic in situ hybridisation (NISH) and polymerase chain reaction (PCR) amplification with primers specific for E6 and E7 regions of the HPV genome. PCR products were analysed by Southern blotting. Immunohistochemical determination of high molecular weight cytokeratin (HMC) and involucrin was also carried out. RESULTS: 18 cases were positive for HPV DNA by PCR and NISH. HPV types 6, 11, 16, and 18 were found. Seven cases were dual positive for different types of HPV. Using NISH, HPV was also found in the squamous cell components and in neighbouring enlarged adenocarcinoma cells. The HMC and involucrin were demonstrated immunohistochemically in the same areas. CONCLUSIONS: HPV DNA was found in a high proportion (78.3%) of adenosquamous carcinomas in Okinawa, a region where HPV has previously been shown to be prevalent in squamous cell carcinoma of the lung. The adenocarcinoma cells adjacent to the squamous cell carcinoma component were enlarged and positive for HPV, HMC, and involucrin. This is thought to indicate the transition from adenocarcinoma to squamous cell carcinoma.  (+info)

(3/10141) The role of alternative splicing of the adhesion molecule, CD44, in lymphoid malignancy.

AIM: To investigate the expression of CD44 isoforms containing variant exon 6 (v6) in a well characterised cohort of patients with non-Hodgkin's lymphoma (NHL) and chronic lymphocytic leukaemia (CLL), and to correlate this with phenotype and disease course. METHODS: Cryostat sections of OCT embedded diagnostic nodal material from NHL patients and cryopreserved mononuclear preparations from CLL patients were used as sources of RNA. After reverse transcription, PCR was carried out with amplimers positioned at either side of the variant exon insertion site to amplify all possible CD44 isoforms. Those isoforms containing v6 were identified after Southern blotting and hybridisation with a radiolabelled oligonucleotide. RESULTS: Of 32 NHL samples analysed, 16 did not express CD44 isoforms containing v6, six expressed an isoform containing exon v6 alone, and 10 expressed v6 long isoforms which contained exon v6 in addition to other variant exons. These data did not correlate with lymphoma classification, disease staging, or the presence or absence of extranodal disease. However, those patients expressing v6 long CD44 isoforms had a worse overall survival than those that did not. The plateau of the survival curves was 50% compared with 82%. No v6 long isoforms were detected in the 21 CLL samples investigated. CONCLUSIONS: The expression of v6 long CD44 isoforms is associated with aggressive disease in NHL, independent of grade, stage, or presence of extranodal disease.  (+info)

(4/10141) Structure of cag pathogenicity island in Japanese Helicobacter pylori isolates.

BACKGROUND: cag pathogenicity island (PAI) is reported to be a major virulence factor of Helicobacter pylori. AIM: To characterise cagA and the cag PAI in Japanese H pylori strains. METHODS: H pylori isolates from Japanese patients were evaluated for CagA by immunoblot, for cagA transcription by northern blot, and for cagA and 13 other cag PAI genes by Southern blot. cagA negative strains from Western countries were also studied. Induction of interleukin-8 secretion from gastric epithelial cells was also investigated. RESULTS: All Japanese strains retained cagA. Fifty nine of 63 (94%) strains had all the cag PAI genes. In the remaining four, cag PAI was partially deleted, lacking cagA transcripts and not producing CagA protein. Details of the PAI of these strains were checked; three lacked cagB to cagQ (cagI) and continuously cagS to cag13 (cagII), and the remaining one lacked cagB to cag8. Western cagA negative strains completely lacked cag PAI including cagA. Nucleotide sequence analysis in one strain in which the cag PAI was partially deleted showed that the partial deletion contained 25 kb of cag PAI and the cagA promoter. Interleukin-8 induction was lower with the cag PAI partial deletion strains than with the intact ones. All Japanese cag PAI deleted strains were derived from patients with non-ulcer dyspepsia, whereas 41 of 59 (70%) CagA-producing strains were from patients with peptic ulcers or gastric cancer (p<0.05). CONCLUSIONS: Most Japanese H pylori strains had the intact cag PAI. However, some lacked most of the cag PAI in spite of the presence of cagA. Thus the presence of the cagA gene is not an invariable marker of cag PAI related virulence in Japanese strains.  (+info)

(5/10141) Comparison of Bombyx mori and Helicoverpa armigera cytoplasmic actin genes provides clues to the evolution of actin genes in insects.

The cytoplasmic actin genes BmA3 and BmA4 of Bombyx mori were found clustered in a single genomic clone in the same orientation. As a similar clustering of the two cytoplasmic actin genes Ha3a and Ha3b also occurs in another lepidopteran, Helicoverpa armigera, we analyzed the sequence of the pair of genes from each species. Due to the high conservation of cytoplasmic actins, the coding sequence of the four genes was easily aligned, allowing the detection of similarities in noncoding exon and intron sequences as well as in flanking sequences. All four genes exhibited a conserved intron inserted in codon 117, an original position not encountered in other species. It can thus be postulated that all of these genes derived from a common ancestral gene carrying this intron after a single event of insertion. The comparison of the four genes revealed that the genes of B. mori and H. armigera are related in two different ways: the coding sequence and the intron that interrupts it are more similar between paralogous genes within each species than between orthologous genes of the two species. In contrast, the other (noncoding) regions exhibited the greatest similarity between a gene of one species and a gene of the other species, defining two pairs of orthologous genes, BmA3 and HaA3a on one hand and BmA4 and HaA3b on the other. However, in each species, the very high similarities of the coding sequence and of the single intron that interrupts it strongly suggest that gene conversion events have homogenized this part of the sequence. As the divergence of the B. mori genes was higher than that of the H. armigera genes, we postulated that the gene conversion occurred earlier in the B. mori lineage. This leads us to hypothesize that gene conversion could also be responsible for the original transfer of the common intron to the second gene copy before the divergence of the B. mori and H. armigera lineages.  (+info)

(6/10141) Molecular cloning and characterization of three cDNAs encoding putative mitogen-activated protein kinase kinases (MAPKKs) in Arabidopsis thaliana.

We isolated three Arabidopsis thaliana cDNA clones (ATMKK3, ATMKK4 and ATMKK5) encoding protein kinases with extensive homology to the mitogen-activated protein kinase kinases (MAPKKs) of various organisms in the catalytic domain. ATMKK3 shows high homology (85% identity) to NPK2, a tobacco MAPKK homologue. ATMKK4 and 5 are closely related to each other (84% identity). Phylogenetic analysis showed that the plant MAPKKs constitute at least three subgroups. The recombinant ATMKK3 and ATMKK4 were expressed as a fusion protein with glutathione S-transferase (GST) in Escherichia coli. Affinity purified GST-ATMKK3 and GST-ATMKK4 proteins contained phosphorylation activity, which shows that both the ATMKK3 and ATMKK4 genes encode functional protein kinases. Northern blot analysis revealed that the ATMKK3 gene expressed in all the organs. The levels of ATMKK4 and 5 mRNAs were relatively higher in steins and leaves than in flowers and roots. We determined the map positions of the ATMKK3, 4 and 5 genes on Arabidopsis chromosomes by RFLP mapping using P1 genomic clones.  (+info)

(7/10141) Cloning, expression, and enzymatic characterization of Pseudomonas aeruginosa topoisomerase IV.

The topoisomerase IV subunit A gene, parC homolog, has been cloned and sequenced from Pseudomonas aeruginosa PAO1, with cDNA encoding the N-terminal region of Escherichia coli parC used as a probe. The homolog and its upstream gene were presumed to be parC and parE through sequence homology with the parC and parE genes of other organisms. The deduced amino acid sequence of ParC and ParE showed 33 and 32% identity with that of the P. aeruginosa DNA gyrase subunits, GyrA and GyrB, respectively, and 69 and 75% identity with that of E. coli ParC and ParE, respectively. The putative ParC and ParE proteins were overexpressed and separately purified by use of a fusion system with a maltose-binding protein, and their enzymatic properties were examined. The reconstituted enzyme had ATP-dependent decatenation activity, which is the main catalytic activity of bacterial topoisomerase IV, and relaxing activities but had no supercoiling activity. So, the cloned genes were identified as P. aeruginosa topoisomerase IV genes. The inhibitory effects of quinolones on the activities of topoisomerase IV and DNA gyrase were compared. The 50% inhibitory concentrations of quinolones for the decatenation activity of topoisomerase IV were from five to eight times higher than those for the supercoiling activities of P. aeruginosa DNA gyrase. These results confirmed that topoisomerase IV is less sensitive to fluoroquinolones than is DNA gyrase and may be a secondary target of new quinolones in wild-type P. aeruginosa.  (+info)

(8/10141) Reduced pyrazinamidase activity and the natural resistance of Mycobacterium kansasii to the antituberculosis drug pyrazinamide.

Pyrazinamide (PZA), an analog of nicotinamide, is a prodrug that requires conversion to the bactericidal compound pyrazinoic acid (POA) by the bacterial pyrazinamidase (PZase) activity of nicotinamidase to show activity against Mycobacterium tuberculosis. Mutations leading to a loss of PZase activity cause PZA resistance in M. tuberculosis. M. kansasii is naturally resistant to PZA and has reduced PZase activity along with an apparently detectable nicotinamidase activity. The role of the reduction in PZase activity in the natural PZA resistance of M. kansasii is unknown. The MICs of PZA and POA for M. kansasii were determined to be 500 and 125 micrograms/ml, respectively. Using [14C]PZA and [14C]nicotinamide, we found that M. kansasii had about 5-fold-less PZase activity and about 25-fold-less nicotinamidase activity than M. tuberculosis. The M. kansasii pncA gene was cloned on a 1.8-kb BamHI DNA fragment, using M. avium pncA probe. Sequence analysis showed that the M. kansasii pncA gene encoded a protein with homology to its counterparts from M. tuberculosis (69.9%), M. avium (65.6%), and Escherichia coli (28.5%). Transformation of naturally PZA-resistant M. bovis BCG with M. kansasii pncA conferred partial PZA susceptibility. Transformation of M. kansasii with M. avium pncA caused functional expression of PZase and high-level susceptibility to PZA, indicating that the natural PZA resistance in M. kansasii results from a reduced PZase activity. Like M. tuberculosis, M. kansasii accumulated POA in the cells at an acidic pH; however, due to its highly active POA efflux pump, the naturally PZA-resistant species M. smegmatis did not. These findings suggest the existence of a weak POA efflux mechanism in M. kansasii.  (+info)



electrophoresis


  • From reliable restriction enzymes to the fast, convenient E-Gel® agarose gel electrophoresis system and BrightStar®-Plus membranes, our products not only help meet your Southern blotting analysis needs, they also speed up the process. (thermofisher.com)
  • Southern Blotting is an elegant technique that can be used to identify the locations of genes and other DNA sequences on restriction fragments separated by gel electrophoresis . (sciencesamhita.com)
  • We've developed an expansive collection of tools and reagents for DNA and RNA gel electrophoresis and blotting. (thermofisher.com)

fragments


  • During hybridization, the labeled probe is incubated with the DNA fragments that are immobilized on the blot under conditions that promote hybridization of complementary sequences. (thermofisher.com)
  • Southern blotting (named after Edwin M. Southern) is one technique used to transfer the denatured, single-stranded fragments from a gel to a thin solid medium. (pinterest.co.uk)

Gene


  • Samples from 40 patients (37 sarcomas and 3 benign mesen chymal tumors) and samples of 15 normal mesenchymal tissues were examined for GLI gene amplification and expression by Southern hybridization, reverse transcription-PCR of tissue RNA, and immunohisto- chemistry, using a new polyclonal GLI antibody developed against an epitope outside of the zinc finger region. (aacrjournals.org)
  • Southern blot analysis confirmed that 13 of these samples were positive for the viral E6 gene. (bmj.com)

hybridization


  • Because ULTRAhyb® buffer maximizes blot sensitivity, for many targets hybridization can typically be performed in just 2 hours. (thermofisher.com)

restriction


  • Obtaining complete fragmentation of your DNA at the intended restriction enzyme sites is a critical step in Southern blot analysis. (thermofisher.com)
  • We offer a wide selection of high-quality restriction enzymes so that you can restriction digest your DNA to help provide clear, unambiguous Southern blot data. (thermofisher.com)

membranes


  • Ensure reliable results in your DNA and RNA blotting experiments through our wide selection of transfer membranes, enzyme conjugates and chemilumescent detection reagents. (perkinelmer.co.uk)

analysis


  • Southern blot analysis reveals information about DNA identity, size, and abundance. (thermofisher.com)
  • We offer one of the most comprehensive portfolios of products for Southern blot analysis. (thermofisher.com)
  • A probe flanking the 3′-end of the vector was used for Southern blot analysis of ES cell clones and tail biopsies of the progeny. (nih.gov)
  • b) Representative Southern blot analysis of tail biopsies of progeny from heterozygous crosses. (nih.gov)

Techniques


  • The mnemonic to remember blotting techniques is "SNoW DRoP" S - Southern - DNA - D N - Northern - RNA - R O. (pinterest.co.uk)

transfer


  • For fast, reproducible transfer, the iBlot® 7-Min Blot offers complete transfer of DNA to the nylon membrane in seven minutes. (thermofisher.com)