Genomic DNA Library Protocols

Analysis of Clones for Exon Trapping and Amplification Amplification and sequencing of the exon-trapped products.

Bacteriophage Lamda Plant Genomic Library Construction Protocol Protocol for bacteriophage lamda plant genomic library construction. Includes: Preparation of the insert DNA; Pilot Sau3A Digestion; Full-scale partial digestions.

Construction of a Yeast Genomic Library Protocol Protocol for the construction of a Yeast genomic library. Includes: Prepare the genomic DNA; Prepare the Library Vector; Ligate the Digested Genomic DNA to the Digested Vector DNA; Prepare Library DNA from Bacteria.

Construction of cDNA Libraries in Eukaryotic Expression Vectors for Construction and Screening This stage summarizes how to construct a cDNA library in a mammalian vector, using commercial kits. These kits are available from several manufacturers and generally include all the required reagents.

Electroporation of the Library into COS-7 Cells for Exon Trapping and Amplification COS-7 cells are transfected with a library of plasmids containing the genomic DNA of interest.

Exon Trapping and Amplification for Construction of the Library Protocol Protocol describes how to construct and amplify a genomic DNA library in a plasmid vector (pSPL3) equipped to express cloned exons in transfected mammalian cells.

Harvesting the mRNA for Exon Trapping and Amplification 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.

Reverse Transcriptase-PCR for Exon Trapping and Amplification Amplified cDNAs containing functional 3' and 5' splice sites are selected by digestion with BstXI and cloned into a Bluescript vector.

Use of PCR for Quality Control of a Genomic DNA Library Protocol Protocol describes the use of PCR to analyze the quality of a human genomic DNA library. It has been optimized for a human genomic DNA library containing fragments between approximately 50 and 500 bp.