Cloning degenerate oligonucleotides

Aloe (Aloe arborescens) is a medicinal plant rich in aromatic polyketides such as pharmaceutically important aloenin (a hexaketide pyrone), aloesin (a heptaketide chromone), and barbaloin (an octaketide anthrone) (Fig. 4a). Pentaketide chromone synthase (PCS) and octaketide synthase (OKS) are novel plant-specific type III PKSs, which were obtained from the aloe plant by RT-PCR cloning using degenerate oligonucleotide primers based on the conserved sequences of known CHS enzymes [30-33]. The deduced amino acid sequences of PCS and OKS are 91% identical (368/403), and show 50-60% identity to those of other CHS superfamily type III PKSs of plant origin OKS shares 60% identity (240/403) with CHS from... 

Christian et al. [69] evaluated the genetic diversity of their dodeca-peptide library by a modified colony hybridization protocol. This protocol allows the discrimination of perfect matches by hybridizing 17 degenerated oligonucleotide probes to the library clones and washing by incrementing the temperature until the perfect matches alone are able to hybridize. The observed number of positive clones for each oligonucleotide probe is correlated with the theorized number. 

Synthesize a specific oligonucleotide that contains all of the sequence possibilities of the aligned sequences. DNA synthesizers are capable of multiple nucleotide additions at each position, which degenerates the position. Oligos that are as much as 1048-fold degenerate have been used successfully in PCR cloning... 

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