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A Magnetic Particle-Based Method for Purifying PCR Products from Solution Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4100

Category: PCR Protocols

The MagneSil system can selectively isolate PCR products that are more than 150-bp long from primers and primer -dimers. The technology can be used with a number of robotic workstations, including Beckman Coulter’s Biomek 2000 and FX Laboratory Automation Workstations. The procedure can also be carried out manually. Typical recovery is more than 80% for a 1-kb product with negligible carryover of primers or nucleotides. - [Read A Magnetic Particle-Based Method for Purifying PCR Products from Solution Protocol]

AFLP For Positional Cloning - http://www-ciwdpb.stanford.edu/publications/methods/aflp.html

Category: Plant Biology Protocols: Plant Genetics Protocols

AFLP was designed as a highly sensitive method for DNA fingerprinting to be used in a variety of fields. We are using this technology to generate DNA based markers for cloning genes involved in phototropic responses in higher plants that have only been identified genetically by mutant phenotype. Protocol includes: Generate polymorphic recombinant F2 (or F3) population; Isolate genomic DNA; Restriction of DNA; Ligation of adapters; Pre-amplification of template DNA; AFLP-PCR; etc. - [Read AFLP For Positional Cloning]

Arabidopsis Thaliana DNA Isolation Procedure - http://wheat.pw.usda.gov/~lazo/methods/lazo/arabdna.html

Category: Plant Biology Protocols: Plant DNA RNA Isolation Protocols

Procedure to isolate DNA from Arabidopsis Thaliana. - [Read Arabidopsis Thaliana DNA Isolation Procedure]

Detect and Quantitate MicroRNA in Laser Capture Microdissection Samples - http://www.ambion.com/techlib/tn/131/6.html

Category: Primary Cell Isolation and Culture Protocols: Laser Capture Microdissection Protocols

Three Ambion kits were used to quantitate specific miRNAs and to detect differential miRNA expression in various mouse brain regions and cell types isolated by laser capture microdissection (LCM). These techniques can be applied to studying miRNA in other species, tissues, and cell types. Includes: Obtain Laser Capture Microdissected Samples; Isolate miRNA from LCM Samples; Quantitate miRNA by qRT-PCR. - [Read Detect and Quantitate MicroRNA in Laser Capture Microdissection Samples]

Detect and Quantitate MicroRNA in Laser Capture Microdissection Samples - http://www.ambion.com/techlib/tn/131/6.html

Category: RNAi Technology Protocols

Information on how detect and quantitate MicroRNA in laser capture microdissection samples. Includes: Obtain Laser Capture Microdissected Samples; Isolate miRNA from LCM Samples; Quantitate miRNA by qRT-PCR; Detection of miRNA in Microdissected Tissue from Mouse Brain by qRT-PCR; Differential Expression of MicroRNA in Whole Brain Tissue Compared to a More Homogeneous Population of Cells. - [Read Detect and Quantitate MicroRNA in Laser Capture Microdissection Samples]

DNA Isolation from Onion Cells - http://www.accessexcellence.org/AE/AEC/AEF/1994/dollard_onionDNA.html

Category: DNA Protocols: DNA Extraction Protocols

Allows students to easily isolate chromosomal DNA from onion cells using the same basic tools and methods that scientists use in the lab. A good introduction to using pipettes. Kate Dollard - [Read DNA Isolation from Onion Cells]

Extraction of Calf Brain Lipids Protocol - http://www.bio.com/protocolstools/protocol.jhtml?id=p1281

Category: Lipid Protocols

Protocol for the extraction of calf brain lipids. Protocol describes a rapid method to isolate lipids from bovine brain tissue using an organic solvent mixture of Chloroform and Methanol. - [Read Extraction of Calf Brain Lipids Protocol]

Harvesting the mRNA for Exon Trapping and Amplification - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3734

Category: DNA Protocols: Genomic DNA Library Protocols

Standard techniques are used here to isolate cytoplasmic RNA from COS cells transfected with a library of recombinant plasmids containing the genomic DNA of interest. - [Read Harvesting the mRNA for Exon Trapping and Amplification]

How to Determine the Species of a Wild-Collected Isolate - http://www.fgsc.net/neurosporaprotocols/How%20to%20determine%20the%20species%20of%20a%20wild-collected%20isolate.pdf

Category: Fungi Protocols: Neurospora Protocols

Information and tips on how to determine the species of a wild-collected isolate. - [Read How to Determine the Species of a Wild-Collected Isolate]

How to isolate DNA; Large Scale - http://www.fgsc.net/neurosporaprotocols/DNA%20prep.pdf

Category: Fungi Protocols: Neurospora Protocols

Information and tips on how to isolate DNA; Large Scale. - [Read How to isolate DNA; Large Scale]

How to Isolate mRNA - http://www.fgsc.net/neurosporaprotocols/HowtoisolatemRNA%20final.pdf

Category: Fungi Protocols: Neurospora Protocols

Information and tips on how to isolate mRNA. - [Read How to Isolate mRNA]

How to Isolate Nondenatured Protein Extracts from Neurospora for Western Analysis - http://www.fgsc.net/neurosporaprotocols/How%20to%20isolate%20nondenatured%20protein%20extracts%20from%20Neurospora%20for%20western%20analysis.pdf

Category: Fungi Protocols: Neurospora Protocols

Information and tips on how to isolate nondenatured protein extracts from Neurospora for western analysis. - [Read How to Isolate Nondenatured Protein Extracts from Neurospora for Western Analysis]

How to Isolate Proteins - http://www.fgsc.net/neurosporaprotocols/How%20to%20isolate%20proteins%20final.pdf

Category: Fungi Protocols: Neurospora Protocols

Information and tips on how to isolate proteins. - [Read How to Isolate Proteins]

How to Isolate Tightly-Coupled Mitochondria from Neurospora crassa mycelium Protocol - http://www.fgsc.net/neurosporaprotocols/How%20to%20Isolate%20Tightly-Coupled%20Mitochondria.pdf

Category: Fungi Protocols: Neurospora Protocols

Protocol on how to Isolate Tightly-Coupled Mitochondria from Neurospora crassa mycelium. A Fast and Reproducible Procedure - [Read How to Isolate Tightly-Coupled Mitochondria from Neurospora crassa mycelium Protocol]

Isolation of Live Bacteria from Adult Insects Protocol - http://www.natureprotocols.com/2006/06/23/isolation_of_live_bacteria_fro.php

Category: Bacterial Protocols

Protocol was developed to isolate Wolbachia from adult Drosophila, but it can be adapted for other insects. In some insects leg removal prior to isolation facilitates hemolymph extrusion. - [Read Isolation of Live Bacteria from Adult Insects Protocol]

Isolation of Mouse Intrahepatic Lymphocytes Protocol - https://catalog.invitrogen.com/index.cfm?fuseaction=iProtocol.unitSectionTree&treeNodeID=9E66370706B026EE3B53EB4B32AD4A09&objectid=6674D486BDDA02EC25183A2F0D9BED25

Category: Mouse Protocols

Provides two protocols that may be used to isolate IHL. One can be used to isolate IHL from multiple livers in parallel, whereas the more elaborate alternate protocol yields more cells per liver but is more appropriately used to recover the IHL from a single liver. - [Read Isolation of Mouse Intrahepatic Lymphocytes Protocol]

Isolation of RNA from C. elegans Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4321

Category: C. elegans Protocols: C. elegans Genetics Protocols

Protocol describes an efficient approach to isolate RNA from Caenorhabditis elegans. - [Read Isolation of RNA from C. elegans Protocol]

Laser-capture Microdissection Protocol - http://www.nature.com/nprot/journal/v1/n2/full/nprot.2006.85.html#a1

Category: Primary Cell Isolation and Culture Protocols: Laser Capture Microdissection Protocols

LCM technology can harvest the cells of interest directly or can isolate specific cells by cutting away unwanted cells to give histologically pure enriched cell populations. A variety of downstream applications exist: DNA genotyping and loss-of-heterozygosity (LOH) analysis, etc. Protocol provides a thorough description of LCM techniques, with an emphasis on tips and troubleshooting advice derived from LCM users. The total time required to carry out this protocol is typically 1–1.5 h. - [Read Laser-capture Microdissection Protocol]

Neural Stem Cell Culture: Neurosphere Generation, Microscopical Analysis and Cryopreservation - http://www.natureprotocols.com/2006/08/25/neural_stem_cell_culture_neuro.php

Category: Stem Cell Protocols: Stem Cell Culture Protocols

Describes the steps in detail to isolate and expand neural stem cells in the form of neurospheres from tissue dissections of the post-natal mouse brain. Procedures for the long term passage of neurospheres and the cryopreservation of neurospheres are also provided. In addition to the guidelines and tips for generating neurosphere cultures, we describe the method to prepare neurospheres for analysis by light microscopy. - [Read Neural Stem Cell Culture: Neurosphere Generation, Microscopical Analysis and Cryopreservation]

Peroxisomes in Saccharomyces cerevisiae - http://mcb.asm.org/cgi/reprint/11/1/510.pdf

Category: Cell Organelle Protocols: Subcellular Fractionation Protocols: Peroxisome Isolation Protocols

To isolate peroxisomes from Saccharomyces cerevisiae of a quality sufficient for in vitro import studies, we optimized the conditions for cell growth and for cell fractionation. Stability of the isolated peroxisomes was monitored by catalase latency and sedimentability of marker enzymes. - [Read Peroxisomes in Saccharomyces cerevisiae]

Rapid Amplification of 3' cDNA Ends 3'-RACE Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3865

Category: PCR Protocols: cDNA Synthesis Protocols

3'-RACE reactions are used to isolate unknown 3' sequences or to map the 3' termini of mRNAs onto a gene sequence. 3'-RACE requires knowledge of a small region of sequence within either the target RNA or a partial clone of cDNA. A population of mRNAs is transcribed into cDNA with an adaptor-primer consisting at its 3' end of a poly(T) tract and at its 5' end of an arbitrary sequence of 30-40 nucleotides. - [Read Rapid Amplification of 3' cDNA Ends 3'-RACE Protocol]

Rapid Isolation of Yeast DNA Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot4039

Category: Yeast Protocols: Yeast Nucleic Acid Protocols: Yeast DNA Isolation Protocols

This method is used to isolate genomic yeast DNA or shuttle plasmids that replicate in both S. cerevisiae and E. coli. The DNA can be used as a template for PCR and for transformation. - [Read Rapid Isolation of Yeast DNA Protocol]

RNA Preparation from Cultured Cells or Tissue Samples Protocol - http://genetics.med.harvard.edu/~cepko/protocol/mike/R2.html

Category: Nucleic Acid Purification Protocols: RNA Isolation Protocols: RNA Isolation fromTissues or Cultured Cells Protocols

Protocol for RNA preparation from cultured cells or tissue samples. This protocol has been used to isolate RNA from relatively small tissue samples. The RNA is clean enough for Rnase protection, cDNA synthesis, and RT-PCR analysis. - [Read RNA Preparation from Cultured Cells or Tissue Samples Protocol]

Separation of Platelets from Whole Blood - http://www.cbrinstitute.org/labs/springer/protocols/azucena_platelets.html

Category: Cell Biology and Cell Culture

This protocol describes how to isolate human platelets from whole blood. Isolated platelets are used for static adhesion assays, for flow chamber assays, flow cytometry measurements, etc. - [Read Separation of Platelets from Whole Blood]

Streaking Lambda Phages Protocol - http://humgen.wustl.edu/hdk_lab_manual/lambda/lambda12.html

Category: Viral Culture and Isolation Protocols: Viral Culture Protocols

Phage are streaked onto a medium to obtain an independent isolate prior to preparing a new lysate. This is done to reduce the likelihood of working with lysates which have become contaminated, and/or have accumulated mutations. - [Read Streaking Lambda Phages Protocol]

Articles (1-6 of 6)

Single Stranded Plasmid DNA Isolation Protocol

A Single Stranded Plasmid DNA Isolation Protocol describing the production and isolation of single-stranded DNA (ssDNA) using bacteriophagemid-containing bacteria and helper phage. Infection of the host cells with helper phage allows for packaging of ssDNA into bacteriophage. The ssDNA can then be isolated from phage particles.

Immobilized Antigen Phage Antibody Selection Protocol

A protocol for the selection of Phage Antibodies using Immobilized Antigen. This method describes the selection of antibodies from bacteriophage antibody libraries that recognize a specific antigen. The phage display library of antibody-displaying phage particles is exposed to antigen attached to a solid substrate (Nunc Immuno™ tubes). The phage particles with affinity for antigen bind to the immobilized antigen and are selected from the library of phage expressing antibodies.

DNA Fragment Purification from Polyacrylamide Gels Protocol

A simple protocol for the purification of DNA fragments from Polyacrylamide Gels.

Picking Transfected ES Cell Clones Protocol

This protocol a protocol on how to generate transfected embryonic stem (ES) cell clones. The previous protocol in this series is the Protocol for Electroporation of ES cells. The next protocol in the series is the Protocol on Disaggregation, Expansion, and Freezing of Transfected ES Clones.

Electrotransformation of BMH 81-17mut S for Isolating Site-Directed dsDNA Mutants

This protocol describes the electroporation of the BMH 81-17 mut S strain that is recommended for tranformation of the site directed mutagenesis of dsDNA (See Protocol on Site-Directed Mutagenesis on Double Stranded DNA). BMH 81-17 mut S are a mismatch repair defective (mut S) Escherichia coli strain. The probability that the two mutations will cosegregate during the first round of DNA replication is increased in this strain.

Vector Design Protocol for Transgenic Mice Generation

The protocol gives general considerations for the design of targeting vectors for transgenic mice. The protocol shares tips in the design of knock-out and knock-in vectors and some of their strategies for producing homologously recombined embryonic stem cells.

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