eDNA

B

Barcode genes

Refers to genes that can be used to identify species. Different regions of DNA mutate at different speeds. Certain regions change at just the right rate to be stable within a species but different between species. These are known as barcode genes (USFWS, 2023). The official barcode gene for animals is Cytochrome Oxidase 1 (COI or cox-1). Other genes used as animal barcodes include 12S, 16S, 18S and Cytochrome-b (cytb).

C

Centrifugation

A method that separates suspensions by spinning them at high speeds. The components are separated according to density and the rotor speed of the centrifuge. This method isn’t used as often in eDNA analysis because it's difficult to centrifuge large sample volumes.

E

Environmental DNA (eDNA) is organismal DNA that is released into the environment. Sources of eDNA include skin, excrement, mucous, saliva, etc. eDNA can be collected in samples (such as from water, sediment, air) which is then used to identify the species through molecular methods.

F

Filtration

DNA in the water is trapped on filter membranes, which is preserved by multiple methods including: lyophilization, molecular grade alcohol, certain buffers or freezing to prevent DNA degradation. This method is the most common technique used in eDNA studies due to the ability to sample large quantities of water.

H

High-Throughput Sequencing

Technology that produces millions of sequences at once. This allows thousands of different organisms from a mixture of species to be sequenced simultaneously so community DNA can be sequenced. Also known as Next-Generation Sequencing (NGS) or parallel sequencing (USFWS, 2023).

I

Inhibited Sample

A sample that contains substances, such as humic acids, that negatively affects PCR detection and increases the chance of false negatives due to assay failure.

Internal Positive Control (IPC)

Endogenous (may already be present) or exogenous (not already present) DNA spiked into the samples at known concentration for simultaneous detection using a multiplexed qPCR assay compatible with the experimental target assay. It's used to help detect any PCR inhibitors or false-negatives during the qPCR reactions as well as confirm DNA amplification.

M

Metabarcoding

Refers to simultaneously sequencing millions of DNA fragments to identify species represented in a pool of DNA. Metabarcoding uses non-specific primers during PCR which is then followed by high-throughput sequencing. This process allows you to identify different species in a single sample, making it ideal for multispecies assessment.

N

Negative Controls

Also referred to as “blanks." Negative controls go through the same process as eDNA samples, but they do not contain target eDNA. They are used to find contamination in the workflow and are used in various stages of the eDNA process (sample collection, DNA extraction, qPCR detection).

P

PCR

Polymerase chain reaction. A process where numerous copies of a specific DNA segment are made through a series of heating and cooling steps. This amplification process can generate millions of copies of a specific DNA target.

Positive Controls

A sample or reaction well that is expected to produce a positive result. It's used to assess reliability of the reagents and processes used in generating eDNA results. Positive controls are used to verify that all the steps worked properly or to assist with troubleshooting if errors occurred during the workflow that could lead to reporting false negatives.

Precipitation

Precipitation (isopropanol/ethanol) is a method to concentrate nucleic acids. The process prevents the DNA from dissolving in water by adding salt and alcohol allowing for visualization of the DNA sample.

Primers

Short synthetic DNA sections that bind to either end of the DNA segment that are then amplified during the PCR process. Primers can be designed to be species-specific or universal. Species-specific primers are designed so that the target species’ DNA can be amplified from a community sample of DNA. Universal primers are more general and allow multiple species’ DNA to be amplified instead of just a single species.

Q

QPCR

Stands for ‘quantitative PCR’, also known as ‘real-time PCR’. Numerous copies of a target sequence are created from template DNA and then detected in real-time. This reaction uses a colored dye that fluoresces during amplification which allows for real-time quantification. If there's no target species present, no fluorescence is detected (USFWS, 2023). Due to its high sensitivity, qPCR is an ideal approach when detecting a single species. qPCR is often used with species-specific primers.

R

Risk Tolerance

How much of an environmental, biodiversity, or financial loss a manager is prepared to handle relative to management objectives or priorities.

S

Sanger Sequencing

Traditional DNA sequencing where each reaction produces a single sequence, so it only works on amplified DNA of a single species (USFWS, 2023). A sequence is a series of nucleotide bases represented by the letters A, T, C & G.

Site

Refers to a specific, physical area where a sample has been collected from.

eDNA - Environmental DNA Glossary - UMESC

B