Rapid polymerase chain reaction amplification using intact bacterial cells.

abstract：:Transfection experiments involving mixed cell cultures pose special problems because of differential uptake and expression of DNA by different cell types. In order to evaluate the expression of transfected DNA in primary cultures of embryonic myocardial cells, we have used immunofluorescence microscopy to analyze expr...

journal_title：BioTechniques

authors： Antin PB,Mar JH,Ordahl CP

更新日期：1988-07-01 00:00:00

abstract：:We report a robust method for studying en masse changes in translation using cDNA arrays. The relative distribution of messenger RNAs (mRNAs) along polysome gradients was monitored by performing cDNA array analysis of each gradient fraction and quantifying the mRNA translational status by regression analysis. Using th...

journal_title：BioTechniques

authors： Mazan-Mamczarz K,Kawai T,Martindale JL,Gorospe M

更新日期：2005-07-01 00:00:00

abstract：:To avoid the time-consuming reprobing process in hybridization analysis, signal-distinguishable probes (32P, 35S or antigenic hapten-labeled DNA) can be added to the same hybridization mixture. After hybridization, an unambiguous result can be obtained by differential or sequential autoradiography. ...

journal_title：BioTechniques

authors： Au LC,Chang KJ,Shih CM,Teh GW

更新日期：1994-04-01 00:00:00

abstract：:Antibiotics are widely useful in medicine, agriculture, and industrial fermentations. However, increasing problems with resistant strains call for restrained use and alternative strategies. Antisense peptide nucleic acids (PNAs) show potent bactericidal effects when targeted against the essential Escherichia coli acpP...

journal_title：BioTechniques

authors： Dryselius R,Nekhotiaeva N,Nielsen PE,Good L

更新日期：2003-11-01 00:00:00

abstract：:Reverse transcription PCR (RT-PCR) represents a sensitive and powerful tool for analyzing RNA. While it has tremendous potential for quantitative applications, a comprehensive knowledge of its technical aspects is required. Successful quantitative RT-PCR involves correction for experimental variations in individual RT...

journal_title：BioTechniques

authors： Freeman WM,Walker SJ,Vrana KE

更新日期：1999-01-01 00:00:00

abstract：:Here we introduce a modified antibody staining method that uses up to 80% less antibody for flow cytometry. We demonstrate this method for the detection of antigens expressed at high, moderate, or low levels in mouse and rat lymphocytes as well as mouse mammary epithelial cells. We obtained reproducibly accurate resul...

journal_title：BioTechniques

authors： Sharma D,Eichelberg MR,Haag JD,Meilahn AL,Muelbl MJ,Schell K,Smits BM,Gould MN

更新日期：2012-07-01 00:00:00

abstract：:Several methods and computer programs have been developed for estimating the size of DNA fragments from gel electrophoresis. However, methods are lacking that may facilitate in optimization of gel conditions. In this article, a computer program called DNAOPT is described, which was developed to assist researchers in t...

journal_title：BioTechniques

authors： Raghava GP

更新日期：1995-02-01 00:00:00

abstract：:A nascent protein remains in a complex with its ribosome and mRNA if the stop codon is deleted from the mRNA. In the same manner, mRNA forms a stable complex with DNA if the transcription termination is blocked. In principle, if both mRNA translation and DNA transcription termination are prevented, the protein should ...

journal_title：BioTechniques

authors： Liu S,Shivashankar GV,Sano T,Libchaber A

更新日期：2000-10-01 00:00:00

abstract：:Current gene synthesis methods often incorporate a PCR amplification step in order to yield final material sufficient for resolution from multiple off-products. These amplification steps can cause stochastic sampling effects that propagate errors in gene synthesis or decrease variability when applied to the constructi...

journal_title：BioTechniques

authors： Lee ZB,Firnhaber C,Clarke J,DeDecker BS

更新日期：2015-09-01 00:00:00

abstract：:The Sequence Analysis Workshop (SAW) is an interactive program for sequence analysis of immunoglobulin variable domains. Sequences for SAW can be obtained from GenBank or from a standard text file. SAW can compare a variable domain to as many as 100 different sequences, calculate the extent of homology, sort the seque...

journal_title：BioTechniques

authors： Elgavish RA,Schroeder HW Jr

更新日期：1993-12-01 00:00:00

abstract：:Loop-mediated isothermal amplification (LAMP), a novel gene amplification method, enables the synthesis of larger amounts of both DNA and a visible byproduct--namely, magnesium pyrophosphate--without thermal cycling. A positive reaction is indicated by the turbidity of the reaction solution or the color change after a...

journal_title：BioTechniques

authors： Goto M,Honda E,Ogura A,Nomoto A,Hanaki K

更新日期：2009-03-01 00:00:00

abstract：:Previous work from this laboratory has shown that immunoporation has the potential for the selective transfection of a range of different animal cells based on their immunological identity. The unique ability of immunoporation to target cells for transfection combined with the high efficiency of transfection and the h...

journal_title：BioTechniques

authors： Tzavelas C,Bildirici L,Rickwood D

更新日期：2004-08-01 00:00:00

abstract：:Human embryonic kidney (HEK293) cells were stably transduced with a retroviral vector containing an expression cassette for a short-lived green fluorescent protein (d2EGFP) and the neomycin resistance gene (Neor). When Neor HEK293 clones were treated with proteasome inhibitors, lactacystin or MG132, an increase in the...

journal_title：BioTechniques

authors： Andreatta C,Nahreini P,Hovland AR,Kumar B,Edwards-Prasad J,Prasad KN

更新日期：2001-03-01 00:00:00

abstract：:We describe a simple and efficient technique that facilitates the amplification of specific mRNA for cloning and sequencing purposes. An mRNA bound to a small piece of membrane filter is used as a template to synthesize complementary DNA. The product of this reaction is then transferred to a new tube and amplified usi...

journal_title：BioTechniques

authors： Ruiz A,Bok D

更新日期：1993-11-01 00:00:00

abstract：:In the quest to move more basic research discoveries into the clinic, the medical community is building training programs to help researchers gain "translational" skills. Sarah Webb explores how PhDs are learning to speak the language of the clinic. ...

journal_title：BioTechniques

authors： Webb SA

更新日期：2014-08-01 00:00:00

abstract：:Gene expression profiles may offer more or additional information than classic morphologic- and histologic-based tumor classification systems. Because the number of tissue samples examined is usually much smaller than the number of genes examined, efficient data reduction and analysis methods are critical. In this rep...

journal_title：BioTechniques

authors： Xiong M,Jin L,Li W,Boerwinkle E

更新日期：2000-12-01 00:00:00

abstract：:A nonimmune phagemid recombinant antibody fragment (rFab) library was generated with a nominal diversity of 1.16 x 10(7) using the QuikChange Multi Site-Directed Mutagenesis kit. Two degenerate primers spanning the third complementarity-determining region (CDR) loops of the antibody fragment light and heavy chain were...

journal_title：BioTechniques

authors： Kelley LL,Momany C

更新日期：2003-10-01 00:00:00

abstract：:Electrophoretic mobility shift analysis (EMSA) is a well-characterized and widely used technique for the analysis of proten-DNA interaction and the analysis of transcription factor combinatorics. Currently implemented EMSA generally involves the time-consuming use of radiolabeled DNA and polyacrylamide gel electrophor...

journal_title：BioTechniques

authors： Clark J,Shevchuk T,Swiderski PM,Dabur R,Crocitto LE,Buryanov YI,Smith SS

更新日期：2003-09-01 00:00:00

abstract：:New cloning vectors have been developed with features to enhance quick allelic exchange in gram-negative bacteria. The conditionally replicative R6K and transfer origins facilitate conjugation and chromosomal integration into a variety of bacterial species, whereas the sacB gene provides counterselection for allelic e...

journal_title：BioTechniques

authors： Mejia-Santana A,Lloyd CJ,Klose KE

更新日期：2021-01-25 00:00:00

abstract：:Vascular targeting agents (VTAs) can be produced by linking antibodies or antibody fragments directed against endothelial cell markers to effector moieties. So far, it has been necessary to produce the components of VTAs (antibody, antibody fragment, linker, and effector) separately and, subsequently, to conjugate the...

journal_title：BioTechniques

authors： Gottstein C,Wels W,Ober B,Thorpe PE

更新日期：2001-01-01 00:00:00

abstract：:Inverse PCR has been used for the recovery of genome regions flanking a known sequence, although its application to metagenome walking is limited due to inefficient amplification from low copy number fragments. Here we present an improved inverse PCR scheme that enables walking of rare fragments in environmental metag...

journal_title：BioTechniques

authors： Uchiyama T,Watanabe K

更新日期：2006-08-01 00:00:00

abstract：:In this special section of BioTechniques, we examine the role of rapid molecular technologies in the detection and identification of agents of infectious disease (ID) and biological weapons (BWs). Besides the threat posed by the global proliferation of BW technologies, there are numerous emerging and reemerging ID age...

journal_title：BioTechniques

authors： Peruski LF Jr,Peruski AH

更新日期：2003-10-01 00:00:00

abstract：:Methods have been developed to rapidly visualize the size distribution of third complementarity-determining regions (CDR3) in immunoglobulin (Ig) and T-cell receptor (TCR) molecules. DNA fragments spanning the Ig or TCR CDR3 are generated by PCR using primers at fixed positions in the variable and constant segments. T...

journal_title：BioTechniques

authors： Raaphorst FM,Tami J,Sanz IE

更新日期：1996-01-01 00:00:00

abstract：:RNA interference (RNAi) is a recent advance that provides the possibility to reduce the expression of specific target genes in cultured mammalian cells with potential applications on a genome-wide scale. However, to achieve this, robust methodologies that allow automated and efficient delivery of small interfering RNA...

journal_title：BioTechniques

authors： Erfle H,Simpson JC,Bastiaens PI,Pepperkok R

更新日期：2004-09-01 00:00:00

abstract：:Recombinant vaccinia viruses (rVACVs) are promising antigen-delivery systems for vaccine development that are also useful as research tools. Two common methods for selection during construction of rVACV clones are (i) co-insertion of drug resistance or reporter protein genes, which requires the use of additional selec...

journal_title：BioTechniques

authors： Liu L,Cooper T,Eldi P,Garcia-Valtanen P,Diener KR,Howley PM,Hayball JD

更新日期：2017-04-01 00:00:00

abstract：:A new procedure is described for the generation of high-titer, helper-free retrovirus vectors employing receptor-mediated, adenovirus-augmented transfection into a standard packaging cell line. Viral titers are increased 30-fold to 100-fold in transiently (> 10(5) infectious units per mL) and stable (> 10(7) infectiou...

journal_title：BioTechniques

authors： von Rüden T,Stingl L,Cotten M,Wagner E,Zatloukal K

更新日期：1995-03-01 00:00:00

abstract：:Stable cell transfection is used for the expression of exogenous genes or cDNAs in eukaryotic cells. Selection of these transfectants requires a dominant selectable marker. A variety of such markers has been identified and is currently in use. However, many of these are not suitable for all cell types or require uniqu...

journal_title：BioTechniques

authors： Baccam M,Huberman E

更新日期：2003-06-01 00:00:00

abstract：:The colony formation assay (CFA) is the gold standard for measuring the effects of cytotoxic agents on cancer cells in vitro; however, in its traditional 6-well format, it is a time-consuming assay, particularly when evaluating combination therapies. In the interest of increased efficiency, the 6-well CFA was converte...

journal_title：BioTechniques

authors： Katz D,Ito E,Lau KS,Mocanu JD,Bastianutto C,Schimmer AD,Liu FF

更新日期：2008-02-01 00:00:00

abstract：:[Formula: see text] Known to be a sturdy weapon in a scientist's arsenal, how has the gene editing tool CRISPR been applied in the fight against COVID-19? ...

journal_title：BioTechniques

authors： Straiton J

更新日期：2020-11-01 00:00:00

abstract：:Site-specific mutagenesis and directional subcloning were accomplished by using the polymerase chain reaction to generate products that can recombine to form circular DNA. This DNA was transfected into E. coli without phosphorylation of primers, restriction enzyme digestion or ligation. Specifically, the polymerase ch...

journal_title：BioTechniques

authors： Jones DH,Howard BH

更新日期：1990-02-01 00:00:00