Question

8. Describe how to evolve an aptamer that only binds Co2+ but not other metals that are often in water (Mg2+, Pb2+, Ca2+...).

Answer 1

• The term “Aptamer” was coined by Andy Ellington. It stems from the Latin terms “aptus,” meaning to fit, and “meros,” meaning part. Aptamers are short, single-stranded DNA or RNA (ssDNA or ssRNA) molecules that can selectively bind to a specific target, including proteins, peptides, carbohydrates, small molecules, toxins, and even live cells.
• Aptamers assume a variety of shapes due to their tendency to form helices and single-stranded loops. They are extremely versatile and bind targets with high selectivity and specificity. Rather than primary sequence, aptamer binding is determined by its tertiary structure. Target recognition and binding involve three dimensional, shape-dependent interactions as well as hydrophobic interactions, base-stacking, and intercalation.
• Aptamers with affinity for a desired target are selected from a large oligonucleotide library through a process called SELEX, which stands for Sequential Evolution of Ligands by Exponential Enrichment. It will produce ssRNA with specific binding site.
• Step 1. Bind oligonucleotide library and discard non-binders.
• Step 2. Elute oligonucleotides that bind desired target.
• Step 3. Perform PCR to amplify eluted binders.
• Repeat steps 1 through 3 using enriched oligonucleotide pool
• Multiple rounds of SELEX (often 6 to 12) are typically performed for completion of SELEX.
• The SELEX process yields ~106 aptamer sequences. Identifying the best candidates is a specialized process employing a variety of analytical techniques at Base Pair.
• As cobalt is naturally occurring and a widely dispersed element, all natural waters contain trace concentrations of this element, sometimes called “background”, i.e. the naturally occurring concentration present in waters due to geologic rather than anthropogenic influence.
• Cobalt, if it is in the form of vitamin B12, is utilised by fauna in the environment. If this low level of cobalt in water was unavailable, a number of the aquatic organisms in these waters would show signs of vitamin B12 deficiency such as not achieving optimal growth or development.Cobalt poisoning is intoxication caused by excessive levels of cobalt in the body.
• Targetting vitamin B12 is best option as to target cobalt ion.