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Agrobacterium-Mediated Transformation in Rice Protocol - http://www.natureprotocols.com/2007/01/18/a_protocol_for_agrobacteriumme.php

Category: Cytogenetics Protocols

Protocol for Agrobacterium-mediated transformation in rice. Agrobacterium-mediated rice transformation method that is applicable to easily cultured varieties in addition to elite japonica varieties that are more difficult to culture. Using this method, transgenic rice plants can be obtained in about 2–3 months with a transformation frequency of 30–50%, both in easily cultured varieties and recalcitrant elite japonica rice. - [Read Agrobacterium-Mediated Transformation in Rice Protocol]

Chromatin Assembly on Template DNA with Transcription Factors and Drosophila S-190 Chromatin Assembl - http://www.bio.com/protocolstools/protocol.jhtml%3Bjsessionid%3D3APITJJIWFIM1R3FQLMCFEWHUWBNSIV0?id=p9061

Category: Drosophila Protocols: Drosophila Genetics Protocols

Protocol described here produces chromatin with regularly spaced nucleosomes having physiological nucleosome repeat lengths; something that can be difficult to achieve with purified components. In addition, chromatin assembled with the S-190 Chromatin Assembly Extract contains the ATP-dependent chromatin remodeling factors necessary for efficient transcription. - [Read Chromatin Assembly on Template DNA with Transcription Factors and Drosophila S-190 Chromatin Assembl]

Darkfield Illumination - http://microscopy.fsu.edu/primer/techniques/darkfield.html

Category: Microscopy Protocols: Optical Microscopy Protocols

The visibility of the faint star light is enormously enhanced against a dark background. This principle is applied in darkfield (also called darkground) microscopy, a simple and popular method for making unstained transparent specimens clearly visible. Such objects often have refractive indices very close in value to that of their surroundings and are difficult to image in conventional brightfield microscopy. - [Read Darkfield Illumination]

Detection of Chromosomal Imbalances Using DOP-PCR and Comparative Genomic Hybridization (CGH) - http://www.roche-applied-science.com/PROD_INF/MANUALS/InSitu/pdf/ISH_95-103.pdf

Category: Cytogenetics Protocols

CGH provides a comprehensive picture of all chromosomal gains and losses within a single experiment. In contrast to banding analysis, CGH requires no preparation of metaphase chromosomes from the cells to be analyzed (which in certain tumor types may be difficult or even impossible). - [Read Detection of Chromosomal Imbalances Using DOP-PCR and Comparative Genomic Hybridization (CGH)]

Enhancing PCR of Very Difficult Regions Protocol - http://www.genome.ou.edu/protocol_enhancing_PCR.html

Category: PCR Protocols: PCR Optimization

Protocol for enhancing PCR of very difficult regions. - [Read Enhancing PCR of Very Difficult Regions Protocol]

Fluorescent Reagents for Live Cell Imaging and Their Introduction into Cells - http://www.cshprotocols.org/cgi/content/abstract/2006/29/pdb.ip20

Category: Microscopy Protocols: Fluorescence Microscopy Protocols

Most biological specimens are relatively transparent, so details of internal and intracellular morphology are difficult to image in untreated living specimens using simple bright-field techniques. Fluorescence microscopy offers greater advantages and possibilities for increasing contrast and determining the specific localization of molecules in cells. Article outlines the three methods most commonly used to introduce an appropriate label into Drosophila tissue without perturbing the process. - [Read Fluorescent Reagents for Live Cell Imaging and Their Introduction into Cells]

Fragmentation of DNA by Sonication Protocol - http://www.cshprotocols.org/cgi/content/abstract/2006/23/pdb.prot4538

Category: Cloning Protocols

Protocol describes a method for DNA fragmentation by sonication. During sonication, DNA samples are subjected to hydrodynamic shearing by exposure to brief periods of sonication. DNA that has been sonicated for excessive periods of time is extremely difficult to clone. - [Read Fragmentation of DNA by Sonication Protocol]

Fundamentals of Growth, Storage, Genetics and Microscopy of Aspergillus nidulans Protocols - http://www.fgsc.net/fgn48/Kaminskyj.htm

Category: Fungi Protocols: Aspergillus Protocols

Compendium of protocols for using Aspergillus nidulans in genetic, molecular, and cell biological investigations, originally written for members of my research group. It also summarizes our common growth media and nutritional supplements, many of which originally appeared elsewhere but now are difficult to locate. Includes: Growth and storage of Aspergillus nidulans conidia; Nutritional supplements for our common auxotrophies; Double mutants; Mitotic mapping - assigning genes to chromosomes; etc - [Read Fundamentals of Growth, Storage, Genetics and Microscopy of Aspergillus nidulans Protocols]

Identification of Positive GATEWAY Expression Clones Protocol - http://www.cshprotocols.org/cgi/content/abstract/2006/29/pdb.prot4647

Category: Bacteria Genetics Protocols

Protocol for the identification of positive GATEWAY expression clones when both the pENTRY and pDEST vectors contain the same marker for bacterial selection. Protocol describes ways in which difficult vector combinations can be used effectively to obtain the appropriate expression clone without having to convert the pENTRY clone or pDEST clone to vectors with compatible antibiotic resistances. - [Read Identification of Positive GATEWAY Expression Clones Protocol]

Ligation Optimization Protocol - http://www.fhcrc.org/science/labs/fero/Protocols/Ligation.html

Category: Cloning Protocols

Protocol can be used to optimize ligation conditions for difficult to clone (e.g. very large) fragments. The principle is to independently characterize the ligation kinetics of the vector and insert DNA fragments and then to combine them in optimal ratios. - [Read Ligation Optimization Protocol]

Measurement of Cell Adhesion Under Static Conditions - http://www.glycotech.com/protocols/Proto8.html

Category: Cell Biology and Cell Culture: Cell Adhesion Protocols

Many proteins and molecules promote cell adhesion including several cell surface carbohydrate binding proteins. Cell adhesion measurements on 96-well microtiter plate format are difficult due to the shear forces generated by washing the wells. The protocol here introduces the use of a liquid-filled wash chamber that separates unbound cells by gravity. This eliminates uncontrolled shear forces and passage of adherent cells through a liquid/air interface. John L. Magnani~GlycoTech Corporation. - [Read Measurement of Cell Adhesion Under Static Conditions]

Optimizing Electrotransfection of Mammalian Cells In Vitro Protocol - http://www.cshprotocols.org/cgi/content/abstract/2006/1/pdb.prot4449

Category: Transfection Protocols

Protocol describes transfection of plasmid DNA into mammalian cell lines using electroporation, a process whereby external application of electric pulses induce cell membrane permeability. Cells in suspension and small volume cells are difficult to transfect, whereas adherent cells and large volume cells are relatively easy. Regardless of cell size or phenotype, transfection efficiency increases with a high concentration of cells in a small volume. - [Read Optimizing Electrotransfection of Mammalian Cells In Vitro Protocol]

Optimizing Electrotransfection of Mammalian Cells In Vitro Protocol - http://www.cshprotocols.org/cgi/content/abstract/2006/1/pdb.prot4449

Category: Transfection Protocols: Electroporation Sonoporation Protocols

Protocol describes transfection of plasmid DNA into mammaliancell lines using electroporation, a process whereby externalapplication of electric pulses induce cell membrane permeability.A number of factors can affect electrotransfection efficiency.In general, cells in suspension and small volume cells are difficultto transfect, whereas adherent cells and large volume cellsare relatively easy. - [Read Optimizing Electrotransfection of Mammalian Cells In Vitro Protocol]

Plant DNA Extraction Protocol - http://wheat.pw.usda.gov/~lazo/methods/lazo/dnaplmy.html

Category: Plant Biology Protocols: Plant DNA RNA Isolation Protocols

A. thaliana has a very small haploid genome and this makes obtaining DNA somewhat difficult. The most notable problem is that DNA is usually contaminated with polysaccharide which inhibit restriction enzymes as well as other DNA modifying enzymes. This problem is most easily solved by using young plants which have not accumulated as much polysaccharide as older plants. The best results are obtained with plants that are two to three weeks post germinated. - [Read Plant DNA Extraction Protocol]

Sequencing Difficult DNA Regions Protocol - http://www.genome.ou.edu/seq_very_difficult_regions.html

Category: DNA Protocols: DNA Sequencing Protocols

Protocol for sequencing difficult regions on DNA. - [Read Sequencing Difficult DNA Regions Protocol]

Articles (1-3 of 3)

Tubulin Polymerization Protocol using GTP and GMPCPP

Tubulin is polymerized into microtubules by incubating tubulin at 37°C with GTP. A nucleation seed is added when the purpose is to assay microtubule elongation. Tubulin can also be polymerized for the purposes of recycling the tubulin or labeling the microtubules with fluorescently labeled tubulin. Based on the protocol by Timothy Mitchison of Harvard University.

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.

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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