Evaluating mixed samples as a source of error in non-invasive genetic studies using microsatellites

Area of Science:

• Wildlife biology
• Conservation genetics
• Molecular ecology

Background:

• Noninvasive genetic sampling (NGS) is increasingly used for wildlife population surveys.
• Limited research exists on potential biases and errors associated with NGS methods.
• Analyzing mixed samples from multiple individuals is a key concern in NGS studies.

Purpose of the Study:

• To evaluate potential errors in analyzing mixed DNA samples using standard noninvasive genetic sampling (NGS) protocols.
• To assess the effectiveness of common genotyping and error-checking procedures in identifying mixed samples.
• To understand the implications of mixed sample errors on wildlife population estimates.

Main Methods:

• Created 128 mixed DNA samples from brown bear (Ursus arctos) hair.
• Genotyped mixed samples at six microsatellite loci using standard NGS protocols.
• Screened samples for errors, including multiple alleles, single-source amplification, and inconsistent electropherograms.

Main Results:

• Initially, five mixed samples yielded acceptable genotypes.
• All mixed samples ultimately showed errors, such as multiple alleles or inconsistent electropherograms.
• Errors included amplification of only one source sample or inconsistent electropherogram patterns.

Conclusions:

• Standard genotyping and error-checking protocols may fail to detect all mixed samples in noninvasive genetic sampling (NGS).
• Mixed sample errors could lead to underestimation of individual numbers in population surveys.
• Researchers must implement rigorous criteria for gel analysis and error detection to mitigate mixed sample biases.