Gene of interest

Many clones must be created to be confident that the genomic library contains the gene of interest. The probability, P, that some number of clones, N, contains a particular fragment representing a fraction, f, of the genome is... 

Because different cell types in eukaryotic organisms express selected subsets of genes, RNA preparations from cells or tissues in which genes of interest are selectively transcribed are enriched for the desired mRNAs. cDNA... 

Recombinant DNA technology can also be used to create modified proteins that are readily purified by affinity chromatography. The gene of interest is Unked to an oligonucleotide sequence that encodes a carboxyl or amino terminal extension to the encoded protein. The... 

Figure 4.3. Univector plasmid fusion system. Cre- loxP mediated site-specific recombination fuses the pUNI and pHOST plasmids at the loxP site. As a result, the gene of interest is placed under the control of the pHOST promoter and fused to any Tag sequences present in the pHOST plasmid.

The reaction differs from excision of the X chromosome because the Entry Clone contains two attL sites and the destination vector contains two attR sites (Hartley et al., 2000). The att sites are mutated to ensure recombination only occurs between attLl and attRl and between attL2 and att.R2. The recombination reaction proceeds through a cointegrate molecule that is resolved to create a destination vector containing the gene of interest with the desired promoter and tag sequences (Fig. 4.6). 

Figure 7.7. Protein microarray on polyvinylidene difluoride filters (PVDF). Proteins are gridded on filter and probed with an antibody to a gene of interest. Protein-ligand interactions could be detected using a labeled ligand as a probe.

Genes of interest can be tagged at either the N or C terminal end. The decision to tag a protein at either the N or the C terminal depends upon the properties of the protein of interest. In our case, all the eukaryotic translation initiation factors were tagged C terminally to allow the endogenous promoter to influence the expression of the tagged protein. 

Figure 3-1 A method to GFP epitope tag a gene of interest. (A) Diagram representing the C and N terminal cassettes. The upstream and downstream primers representing 55 nucleotides upstream and downstream (but not including the stop codon) are depicted by I H and respectively, and represent the amplification sequence com-...

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