pairs Protocols

Category: PCR Protocols: PCR Cloning Protocols

Pairs of oligonucleotide primers used in PCR are often designed with restriction sites in their 5' regions.  In many cases, the sites are different in the two primers.  In this case, amplification generates a target fragment whose termini now carry new restriction sites that can be used for directional cloning into plasmid vectors.  The purified fragment and the vector are digested with the appropriate restriction enzymes, ligated together, and transformed into E. coli. - [Read Cloning PCR Products by Addition of Restriction Sites to the Termini of Amplified DNA Protocol]

Category: C. elegans Protocols: C. elegans Fixing Protocols

Bouin’s fixative is a particularly good choice for worms because it penetrates dense tissues well and is extremely good for fixing antigens. Like all strong fixatives, however, it is unsuitable for some antibody-antigen pairs. In such cases, the length of time in the Bouin’s fixative can be shortened, or paraformaldehyde fixation can be used instead. - [Read Fixing Caenorhabditis elegans in Bouin’s Fixative Protocol]

Category: Tetrahymena Protozoa Protocols

Mature Tetrahymena cells of opposite mating types are starved under appropriate salt conditions. The mating types are then combined to costimulate through cell-cell interaction. Loose pairs and then firm, irreversible pairs of cells of opposite mating types form. This method consistently results in a high percentage of pairing (usually greater than 80%) and good synchrony. - [Read Induction of Conjugation in Tetrahymena Protocol]

Category: Genetics and Genomics: Genotyping Protocols: PCR Genotyping

Protocol for PCR genotyping from tail DNA. This protocol works well for a variety of genes and primer pairs including Tg and KO alleles. Oligonucleotide melting temperatures between 60° and 65° seem to work well. - [Read PCR Genotyping from Tail DNA Protocol]

Category: Genetics and Genomics: Genotyping Protocols: PCR Genotyping

Primer pairs will amplify sequences present as a single copy in the mouse genome with the Universal Genotyping Protocol. Includes: b-Galactosidase (LacZ); cre-recombinase; CFP; diphtheria toxin; dsRED; Fabpi-200; Fabpi-500; flp recombinase; GFP/BFP/YFP; human growth hormone (complete); human growth hormone (transcriptional stop); luciferase (click-beetle); luciferase (firefly); neomycin phosphotransferase; SRY (male-specific); tTA (tet-on). - [Read PCR Genotyping Primer Pairs Protocols]