Production of stably transfected cell lines using immunoporation.

abstract：:The high-order structure of human chromosomes is an important biological question that is still under investigation. Studies have been done on imaging human mitotic chromosomes using mostly 2-D microscopy methods. To image micron-sized human chromosomes in 3-D, we developed a procedure for preparing samples for serial...

journal_title：BioTechniques

authors： Yusuf M,Chen B,Hashimoto T,Estandarte AK,Thompson G,Robinson I

更新日期：2014-12-01 00:00:00

abstract：:Ditag genome scanning (DGS) uses next-generation DNA sequencing to sequence the ends of ditag fragments produced by restriction enzymes. These sequences are compared to known genome sequences to determine their structure. In order to use DGS for large-scale genome structural studies, we have substantially revised the ...

journal_title：BioTechniques

authors： Jung YC,Xu J,Chen J,Kim Y,Winchester D,Wang SM

更新日期：2009-11-01 00:00:00

abstract：:We observed the presence of contaminating NADH oxidation activity in maltose binding protein (MBP) fusion proteins expressed in Escherichia coli and purified using conventional amylose resin-based affinity chromatography. This contaminating NADH oxidation activity was detectable with at least four different enzymes fr...

journal_title：BioTechniques

authors： Guo F,Zhu G

更新日期：2012-04-01 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：:High stabilities of reporter proteins and their messenger RNAs (mRNAs) interfere with the detection of rapid transient changes in gene expression, such as transcriptional blocks posed by genotoxic DNA lesions. We have modified a green fluorescent protein (GFP) gene within the episomal pMARS vector by addition of a fra...

journal_title：BioTechniques

authors： Kitsera N,Khobta A,Epe B

更新日期：2007-08-01 00:00:00

abstract：:We describe an expeditious method for the isolation of cDNA clones utilizing PCR-based amplification of target sequences from cDNA libraries. This method is rapid, less labor-intensive and inexpensive when compared with screening libraries with radiolabeled probes. This method can be applied to isolate multiple member...

journal_title：BioTechniques

authors： Isola NR,Harn HJ,Cooper DL

更新日期：1991-11-01 00:00:00

abstract：:We describe the application of simple cloning by prolonged overlap extension for multiple site-directed mutagenesis in the same plasmid. We show that it is possible to use this technique with very short PCR templates. The technique is ideally suited for the generation of longer donor DNA sequences for CRISPR/Cas9-medi...

journal_title：BioTechniques

authors： Hejlesen R,Füchtbauer EM

更新日期：2020-06-01 00:00:00

abstract：:The systematic preparation of synthetic peptide combinatorial libraries (SPCLs), each composed of tens of millions of peptides that can be screened in existing diagnostically or pharmacologically relevant in vitro assay systems, is reviewed. The identification of optimal peptide sequences has been achieved through the...

journal_title：BioTechniques

authors： Houghten RA,Appel JR,Blondelle SE,Cuervo JH,Dooley CT,Pinilla C

更新日期：1992-09-01 00:00:00

abstract：:We report a modification of a liquid hybridization method that serves as a rapid, sensitive alternative to Southern hybridization for the analysis of polymerase chain reaction (PCR) products. An aliquot of the completed PCR is mixed with an internally nested, end-labeled oligonucleotide probe, and one cycle of PCR is ...

journal_title：BioTechniques

authors： Parker JD,Burmer GC

更新日期：1991-01-01 00:00:00

abstract：:A streamlined protocol is described that allows high sensitivity antigen detection by Western blotting in a single day. The choice of membrane blotting matrix, as well as blocking reagents, has been optimized in order to allow rapid development of the blot with chemiluminescent reagents. The entire process, from gel t...

journal_title：BioTechniques

authors： Klapper A,MacKay B,Resh MD

更新日期：1992-05-01 00:00:00

abstract：:There is an enormous initiative to establish the genetic basis for disorders of brain function. Unfortunately, genetic intervention is not accomplished easily in the nervous system. One strategy is to engineer and deliver to neurons specialized viral vectors that carry a gene (or genes) of interest, thereby exploiting...

journal_title：BioTechniques

authors： Neve RL,Neve KA,Nestler EJ,Carlezon WA Jr

更新日期：2005-09-01 00:00:00

abstract：:DNA damage in the form of abasic sites, chemically altered nucleotides, and strand fragmentation is the foremost limitation in obtaining genetic information from many ancient samples. Upon cell death, DNA continues to endure various chemical attacks such as hydrolysis and oxidation, but repair pathways found in vivo n...

journal_title：BioTechniques

authors： Mouttham N,Klunk J,Kuch M,Fourney R,Poinar H

更新日期：2015-07-01 00:00:00

abstract：:The use of reverse transcription (RT) PCR for relative quantitation of gene transcripts relies on the reproducibility of the individual RT, PCR and product measurement steps. Semi-competitive RT-PCR (RT-cPCR) uses an internal competitor template in the PCR step to improve quantitation. We have surveyed the reproducibi...

journal_title：BioTechniques

authors： Hall LL,Bicknell GR,Primrose L,Pringle JH,Shaw JA,Furness PN

更新日期：1998-04-01 00:00:00

abstract：:A fully automated nucleic acid analysis system is described, which offers positive sample identification, improved sensitivity and reduced user interaction compared to conventional techniques. The system relies on the sequence-specific capture of DNA onto solid-phase particles, confirming product identity without the ...

journal_title：BioTechniques

authors： Brown A,Akinsanya AA,Barker SJ,Brophy M,Dobb AK,Doyle SM,Hudson IR,Minter SJ,Wraith MJ,Oultram JD

更新日期：1999-07-01 00:00:00

abstract：:Cycling probe technology (CPT) is a simple signal amplification method for the detection of specific target DNA sequences. CPT uses a chimeric DNA-RNA-DNA probe that is cut by RNase H when bound to its complementary target sequence. In this study, a hybridization assay was developed to detect biotinylated CPT products...

journal_title：BioTechniques

authors： Warnon S,Zammatteo N,Alexandre I,Hans C,Remacle J

更新日期：2000-06-01 00:00:00

abstract：:The oligonucleotide ligation assay (OLA) was adapted to the genotyping of the N-arylamine-acetyltransferase (NAT2) gene. This assay allows the use of 96-well microplates and robotic workstations for high sample throughput. We found this assay to be accurate, efficient and reliable. Another advantage for epidemiologica...

journal_title：BioTechniques

authors： Bigler J,Chen C,Potter JD

更新日期：1997-04-01 00:00:00

abstract：:Rapid identification of viruses is needed to monitor the blood supply for emerging threats. Here we present a method that meets these criteria and allows for the shotgun sequencing of novel, uncultured DNA viruses directly from human blood. This method employs selection based on the physical properties of viruses comb...

journal_title：BioTechniques

authors： Breitbart M,Rohwer F

更新日期：2005-11-01 00:00:00

abstract：:A rapid and inexpensive method for isolating bacterial DNA for use in PCR is described. The method is based on the guanidinium thiocyanate (GuSCN)-lysis method of Boom et al. (J. Clin. Microbiol. 28:495-503, 1990) and enables a multiple of 96 samples to be prepared in only one hour. We use Multiscreen plates and a vac...

journal_title：BioTechniques

authors： Reek FH,Smits MA,Kamp EM,Smith HE

更新日期：1995-08-01 00:00:00

abstract：:High-throughput genotyping technologies such as DNA pooling and DNA microarrays mean that whole-genome screens are now practical for complex disease gene discovery using association studies. Because it is currently impractical to use all available markers, a subset is typically selected on the basis of required satura...

journal_title：BioTechniques

authors： Simpson CL,Hansen VK,Sham PC,Collins A,Powell JF,Al-Chalabi A

更新日期：2004-12-01 00:00:00

abstract：:A method for the isolation of total cytoplasmic RNA and high molecular weight DNA from the same cells is described. Cells are gently lysed with NP40 in the presence of vanadyl ribonucleoside complex and the nuclei pelleted by centrifugation. RNA is purified by phenol/CHCl3 extraction of the lysate supernatant followed...

journal_title：BioTechniques

authors： Nicolaides NC,Stoeckert CJ Jr

更新日期：1990-02-01 00:00:00

abstract：:We have demonstrated that efficient polymerase chain reaction amplifications from chromosomal DNA can be carried out using whole bacterial cells as the starting material. Cells from the liquid or solid cultures can be used directly, without any pre-treatment, thus eliminating the need for DNA isolation. ...

journal_title：BioTechniques

authors： Joshi AK,Baichwal V,Ames GF

更新日期：1991-01-01 00:00:00

abstract：:Duracryl is a mechanically strong and elastic acrylamide-based matrix, useful for a wide variety of electrophoretic applications. The matrix is stable as a refrigerated solution for one year. Upon addition of appropriate catalysts, Duracryl forms a polymer-reinforced polyacrylamide gel matrix suitable for electrophore...

journal_title：BioTechniques

authors： Patton WF,Lopez MF,Barry P,Skea WM

更新日期：1992-04-01 00:00:00

abstract：:Conventional genomic DNA (gDNA) extraction methods can take hours to complete, may require fume hoods and represent the most time-consuming step in many gDNA-based molecular assays. We systematically optimized a bead bashing-based (BBB) approach for rapid gDNA extraction without the need for a fume hood. Human tissue ...

journal_title：BioTechniques

authors： Wei S,Levy B,Hoffman N,Cujar C,Rodney-Sandy R,Wapner R,D'Alton M,Williams Z

更新日期：2020-05-01 00:00:00

abstract：:Reducing the scale of biochemical reactions is becoming commonplace. Examples include the screening of large libraries of chemical compounds or gene sequences. These applications demand the ability to transfer sub-microliter volumes of fluid. To this end, we have modified a Hamilton MICROLAB 2200 with high-speed solen...

journal_title：BioTechniques

authors： Hicks JS,Harker BW,Beattie KL,Doktycz MJ

更新日期：2001-04-01 00:00:00

abstract：:Here we describe a convenient method to generate homologous recombinant baculoviral genomes in E. coli. The recombination takes place with the aid of recombination enzymes provided by the phage lambda Red system between a bacmid (a baculoviral genome that can replicate in bacteria) and a linear fragment. Proof of conc...

journal_title：BioTechniques

authors： Hou S,Chen X,Wang H,Tao M,Hu Z

更新日期：2002-04-01 00:00:00

abstract：:In this study, we have developed an air-interface culture system in which guinea pig tracheobronchial epithelial (GPTE) cells rapidly form tight monolayers. Enzymatically isolated GPTE cells were plated on collagen-treated polycarbonate microporous cell culture inserts at a density of 10(6) cells/cm2 (day 0). Bioelect...

journal_title：BioTechniques

authors： Robison TW,Dorio RJ,Kim KJ

更新日期：1993-09-01 00:00:00

abstract：:Francisella tularensis is one of the most deadly bacterial agents, yet most of the genetic determinants of pathogenesis are still unknown. We have developed an efficient targeted mutagenesis strategy in the model organism F. tularensis subsp. novicida by utilizing universal priming of optimized antibiotic resistance c...

journal_title：BioTechniques

authors： Liu J,Zogaj X,Barker JR,Klose KE

更新日期：2007-10-01 00:00:00

abstract：:Single-cell genomics (SCG) is a recently developed tool to study the genomes of unculturable bacterial species. SCG relies on multiple-strand displacement amplification (MDA), PCR, and next-generation sequencing (NGS); however, obtaining sufficient amounts of high-quality DNA from samples is a major challenge when per...

journal_title：BioTechniques

authors： He J,Du S,Tan X,Arefin A,Han CS

更新日期：2016-03-01 00:00:00

abstract：:High-throughput approaches for gene cloning and expression require the development of new nonstandard tools for molecular biologists and biochemists. We introduce a Web-based tool to design primers specifically for the generation of expression clones for both laboratory-scale and high-throughput projects. The applicat...

journal_title：BioTechniques

authors： Yoon JR,Laible PD,Gu M,Scott HN,Collart FR

更新日期：2002-12-01 00:00:00

abstract：:We describe a simple and efficient system for epitope mapping by cloning random gene fragments into a specially designed gIIIp-based phage display vector. DNA encoding the antigen of interest is PCR-amplified and partially digested with DNaseI to generate 50-150-bp-long fragments, which are polished with T4 DNA polyme...

journal_title：BioTechniques

authors： Gupta S,Arora K,Sampath A,Khurana S,Singh SS,Gupta A,Chaudhary VK

更新日期：1999-08-01 00:00:00