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Annealing siRNAs to Produce siRNA Duplexes Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4340

Category: RNAi Technology Protocols

Protocol describes a procedure to anneal sense and antisense siRNA strands to form a duplex. The duplexes can then be transfected into cultured cells. - [Read Annealing siRNAs to Produce siRNA Duplexes Protocol]

Construction of the Sequencing Library Protocol - http://sequence-www.stanford.edu/protocols/library.html

Category: DNA Protocols: DNA Sequencing Protocols

Protocol first describes the vector preparation and, then, describes the insert preparation. Vital to have an excellent vector in order to produce a sequencing library. Protocol employs the male-specific coliphage M13 as the sequencing vector. M13 is a filamentous phage with a single-stranded, circular genome. M13 is widely used as a vector because many versions are available commercially and because M13 has certain advantages. - [Read Construction of the Sequencing Library Protocol]

Detection of Autoantibodies with Self-Assembling Radiolabeled Antigen Tetramers Protocol - http://www.natureprotocols.com/2007/01/29/detection_of_autoantibodies_wi.php

Category: Immunology

Protocol for detection of autoantibodies with self-assembling radiolabeled antigen tetramers. Details how to produce radiolabeled antigen-streptavidin tetramers for detection of antibodies by immunoprecipitation. Optionally, the antigen tetramers can be denatured to compare responses to folded and unfolded antigen in the same system. This technique can be applied to a large or small number of samples, and a given sample can be simultaneously assayed with multiple antigens. - [Read Detection of Autoantibodies with Self-Assembling Radiolabeled Antigen Tetramers Protocol]

Fabrication Protocol for DNA Microarrays - http://research.nhgri.nih.gov/microarray/fabrication.shtml

Category: DNA Microarray Protocols

This protocol describes the steps required to produce a cDNA microarray. Gene-specific DNA is produced by PCR amplification of purified template plasmid DNAs from cloned ESTs. The PCR product is purified by ethanol precipitation, thoroughly resuspended in - [Read Fabrication Protocol for DNA Microarrays]

Growth and Purification of 25-100 ug Lambda Clone DNA Protocol - http://humgen.wustl.edu/hdk_lab_manual/lambda/lambda13.html

Category: Viral Culture and Isolation Protocols: Viral Culture Protocols

There are essentially three parts to this protocol: 1. growth of at least 5x10e8 pfu phage to provide an inoculum growth of a larger liquid lysate that will produce about 5x10e12 pfu; 2. concentration and purification of the phage, and; 3. DNA preparation. - [Read Growth and Purification of 25-100 ug Lambda Clone DNA Protocol]

His Tag Nickel Affinity Chromatography Protocol PDF - http://www.mnstate.edu/provost/His_TagNiChromatogProtocol.pdf

Category: Chromatography Protocols

His Tag Nickel Affinity Chromatography Protocol PDF. The Wallert and Provost Lab. Theory and Introduction: Ni-Affinity Chromatography uses the ability of His to bind nickel. Six histadine amino acids at the end of a protein (either N or C terminus) is known as a 6X His tag. Nickel is bound to an agarose bead by chelation using nitroloacetic acid (NTA) beads. Several companies produce these beads as His Tagged proteins are some of the most used affinity tags in todayâ€™s market. - [Read His Tag Nickel Affinity Chromatography Protocol PDF]

Home-made Taq Polymerase Purification - http://www.nottingham.ac.uk/~mbzspd/methods/Home_made_Taq_polyme.html

Category: PCR Protocols

PCR polymerase costs can be high. If you are willing to work, you can produce bacteria containing the clone. It appears to produce lots of Taq and is quite stable. The proceedure takes 4 days start to (15 000 units of Taq) finish. The Taq also appears ver - [Read Home-made Taq Polymerase Purification]

Imaging Actin in Tissue Slices from Transgenic Mouse Brain Protocol - http://www.cshprotocols.org/cgi/content/extract/2007/3/pdb.ip28

Category: Cytoskeleton Protocols: Actin Myosin Protocols

The same GFP-tagged actin construct used in cell transfection experiments has been used to produce transgenic mice. Transgenic animals allow the imaging of brain tissue in the intact animal, as acutely cut slices or as organotypic slice cultures. They also serve as a source of cells for imaging neurons at high resolution in dispersed low-density cell culture. In contrast to cells transfected in culture, where the level of actin-GFP expression in neurons varies considerably, transgenic mice... - [Read Imaging Actin in Tissue Slices from Transgenic Mouse Brain Protocol]

Injecting Blastocysts Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/21/pdb.prot4418

Category: Stem Cell Protocols: Stem Cell Injection Protocols

This protocol describes a method for injecting mouse blastocysts with embryonic stem (ES) cells to produce chimeras. - [Read Injecting Blastocysts Protocol]

Introduction of Double-Stranded RNA in C. elegans by Soaking Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4316

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

Double-stranded RNA (dsRNA) can be introduced into Caenorhabditis elegans by soaking the animals in a solution of dsRNA. Alternative methods are dsRNA injection (see Introduction of Double-stranded RNA in C. elegans by Injection) and feeding the animals with bacteria that produce dsRNA. - [Read Introduction of Double-Stranded RNA in C. elegans by Soaking Protocol]

Introduction of Double-Stranded RNA in C. elegans by Soaking Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4316

Category: RNAi Technology Protocols: RNAi C. elegans Protocols

Double-stranded RNA (dsRNA) can be introduced into Caenorhabditis elegans by soaking the animals in a solution of dsRNA. Alternative methods are dsRNA injection and feeding the animals with bacteria that produce dsRNA. - [Read Introduction of Double-Stranded RNA in C. elegans by Soaking Protocol]

Large-Scale Preparation of Agarose Plugs of Yeast DNA Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/16/pdb.prot4392

Category: Yeast Protocols: Yeast Genetics Protocols: YACs Protocols

Protocol describes the use of agarose plugs for isolation of yeast artificial chromosome (YAC) DNA. The DNA can then be run on a pulsed-field gel and used for microinjection to produce transgenic mice. - [Read Large-Scale Preparation of Agarose Plugs of Yeast DNA Protocol]

Looking Inside Cells and Tissues by Optical Sectioning with a Confocal Laser Scanning Microscope - http://www.mih.unibas.ch/Booklet/Booklet96/Chapter1/Chapter1.html

Category: Microscopy Protocols: Confocal Microscopy Protocols

Confocal laser scanning microscopy (CLSM) is a relatively new light microscopical imaging technique which has found wide applications in the biological sciences. The primary value of the CLSM to the biologist is its ability to produce optical sections through a 3-D specimen-e.g., an entire cell or a piece of tissue - that, to a good approximation, contain information from only one focal plane. Article includes principle and applications of confocal laser scanning microscope. - [Read Looking Inside Cells and Tissues by Optical Sectioning with a Confocal Laser Scanning Microscope]

Microinjection of Mouse Zygotes Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/16/pdb.prot4397

Category: Microinjection Protocols: Microinjection of DNA Protocols

This protocol describes a method for microinjection of mouse zygotes to produce transgenic mice. - [Read Microinjection of Mouse Zygotes Protocol]

Microinjection Setup Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/16/pdb.prot4396

Category: Microinjection Protocols: Microinjection Setup Protocols

This protocol describes the microinjection setup for injecting mouse zygotes to produce transgenic mice. - [Read Microinjection Setup Protocol]

Monoclonal Antibody Production Fusion Protocol - http://www.icp.ucl.ac.be/mexp/vipg_fus.html

Category: Antibody Protocols: Antibody Production Protocols

Protocol for monoclonal antibody production fusion. Fusion is an important procedure to produce monoclonal antibodies (MoAb). - [Read Monoclonal Antibody Production Fusion Protocol]

Preparation of a Highly Efficient Transcription Extract from Drosophila Embryos Protocol - http://www.bio.com/protocolstools/protocol.jhtml%3Bjsessionid%3D3APITJJIWFIM1R3FQLMCFEWHUWBNSIV0?id=p9052

Category: Drosophila Protocols: Drosophila Genetics Protocols

Protocol describes how to produce a soluble nuclear extract rich in basal pol II transcription factors from Drosophila embryos. This is a cell-free extract that contains all the necessary transcription factors and is capable of accurate initiation of transcription by RNA polymerase II but is deficient in core histones and histone H1. - [Read Preparation of a Highly Efficient Transcription Extract from Drosophila Embryos Protocol]

Preparation of Lysates Containing Fusion Proteins Encoded by Bacteriophage {lambda}: Lytic Infection - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3771

Category: Viral Culture and Isolation Protocols: Viral Culture Protocols

This rapid method is used to screen bacteriophage {lambda}gt11 clones for the production of immunodetectable fusion proteins. After optimizng the conditions of infection and induction, the method can be used to produce preparative amounts of a fusion protein. - [Read Preparation of Lysates Containing Fusion Proteins Encoded by Bacteriophage {lambda}: Lytic Infection]

Preparing and Transforming Spheroplasts of Neurospora crassa Protocol - http://www.fgsc.net/neurosporaprotocols/How%20to%20Prepare%20Spheroplasts.pdf

Category: Fungi Protocols: Neurospora Protocols

Procedure for preparing and transforming spheroplasts of Neurospora crassa. The procedure is designed to produce a large number of spheroplasts to be stored frozen at -80°. - [Read Preparing and Transforming Spheroplasts of Neurospora crassa Protocol]

Preparing Embryonic Stem (ES) Cells for Aggregation Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/21/pdb.prot4419

Category: Stem Cell Protocols

Ths protocol describes the preparation of ES cells, which can then be aggregated with diploid or tetraploid embryos to produce aggregation chimeras. - [Read Preparing Embryonic Stem (ES) Cells for Aggregation Protocol]

reparation of Copy Standards for Southern Blot Copy Number Determination - http://www.med.umich.edu/tamc/spike.html

Category: Transgenic Mouse Protocols: Mouse Genotyping Protocols

PCR screens must be designed to detect transgene DNA at the single copy level.Copy standards are prepared by mixing non-transgenic tail DNA with a known amount of transgene DNA to produce transgene copy standards. University of Michigan Transgenic Animal - [Read reparation of Copy Standards for Southern Blot Copy Number Determination]

Setting Up Microdrop Cultures Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4353

Category: Stem Cell Protocols: Stem Cell Culture Protocols

Protocol describes how to set up microdrop cultures to produce embryos which can then be used for making chimeras. The microdrop culture should be set up several hours to 1 day before the experiment to permit temperature and gas equilibration. - [Read Setting Up Microdrop Cultures Protocol]

Simple, Reliable Steps for DNA Fragment Isolation and Purification Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/16/pdb.prot4389

Category: Mouse Protocols: Mouse Genetics Protocols

Protocol describes a simple procedure for DNA fragment isolation and purification. The resulting DNA fragment can then. be used for microinjection to produce transgenic mice - [Read Simple, Reliable Steps for DNA Fragment Isolation and Purification Protocol]

Articles (1-4 of 4)

Genomic DNA Labeling of Microarrays Protocol

DNA microarrays are an ordered arrangement of DNA molecules complementary to genes of interest that are "spotted" by robotic equipment onto a glass slide substrate. The expression of genes in cells can be monitored with microarrays by preparing cDNA from the mRNA of cells of interest and measuring the hybridization to the microarray. This protocol describes the labeling of genomic DNA for use as a probe for hybridization to the cDNA spotted on the array.

Coupling Cysteine Containing Peptides to Carrier Protein for Immunization and Polyclonal Antibody

This protocol is used in the production of polyclonal antibodies that recognize a peptide sequence of interest.

DNA Ligation Protocol

The DNA Ligation protocol described here contains the steps required to join together using ligase enzyme both plasmid DNA and insert DNA fragments in order to create a new plasmid. This new ligated plasmid can be transformed after into competent bacteria to produce DNA for mini, midi or maxi-prep isolation.

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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