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Aiming the isolation and molecular characterization of the gene for the d polypeptide chain of giant extracellular hemoglobin of Glossoscolex paulistus (HbGp), we present the design of oligonucleotides to DNA hybridization procedures. From a described amino acid sequence, we performed reverse translation, with an algorithm, which searched for regions with maximum density of non-degenerate codons, focusing minimization of the number of putative oligonucleotide probes. Fourteen different partial oligopeptide sequences six, seven or eight residues long resulted in 64, 256 and 512 possible coding sequences, respectively. Based on the similarity with d chain of the hemoglobin from Lumbricus terrestris (HbLt), we ranked the resulting sequences. This narrowed the candidate sequences to 2, 20 and 24 possible probes 18, 21 and 24 nucleotides long, respectively, from selecting the highest scoring sequences. We filtered the set of putative probes based on the relative frequency of codon usage from L. terrestris. Sequences with the higher scores were chosen, favoring shorter sequences. Thus, five sequences targeting two different regions and possible primer pairs for PCR with the HbGp d chain gene were obtained. These oligonucleotides can be employed in hybridization and PCR essays in G. paulistus genomic DNA.

Bioinformatical Development of Oligonucleotides for the d Chain Gene of the Giant Extracellular Hemoglobin of Glossoscolex Paulistus