Skip navigation
Go to home page > News > Our blog > Using PCR to assess the health of Galapagos giant tortoises
Galapagos giant tortoise (Santa Cruz)
03/08/2022 Genetics Women in science

Using PCR to assess the health of Galapagos giant tortoises

Gislayne, a biotechnologist from Galapagos, explores the use of PCR in work to assess the health of Endangered giant tortoises.

Photograph of Gislayne Mendoza Alcívar

Gislayne Mendoza Alcívar

Gislayne Mendoza Alcívar is a Galapagos-born biotechnologist at the Charles Darwin Foundation, whose work with the Galapagos Tortoise Movement Ecology Programme has been funded by GCT.

Gislayne Mendoza is a biotechnologist from Galapagos working with the Charles Darwin Foundation (CDF) and funded by Galapagos Conservation Trust (GCT). In this blog, she explores the use of a specific scientific technique called Polymerase Chain Reaction technique (PCR) in fascinating work to assess the health of Endangered giant tortoises as part of the Galapagos Tortoise Movement Ecology Programme (GTMEP).

Ainoa Nieto preparing materials to collect biological samples from Alcedo Volcano giant tortoises Photo
Ainoa Nieto preparing materials to collect biological samples from Alcedo Volcano giant tortoises © Juan Manuel García, CDF

In the warm month of May 2021, I started working at the Charles Darwin Research Station with GTMEP. As a Galapagueña, this has been a great opportunity to return home after finishing my undergraduate degree in Biotechnology Engineering at the Universidad San Francisco de Quito and to contribute to the study and conservation of our iconic giant tortoises.

I started working with Dr Ainoa Nieto, Principal Investigator of GTMEP. Under her mentorship, we continued working in the molecular biology area, which had been stopped during the COVID-19 pandemic. My job was to detect and analyse specific viruses in Galapagos tortoises using the Polymerase Chain Reaction technique (PCR). But … what actually is a PCR?

PCR is a technique that has been used since Dr Kary Mullis discovered it in 1990, but it has become world-renowned in the wake of the recent pandemic. Despite the fact that almost everyone on the planet has heard this word in the last two years, or even recently undergone a PCR, it is very likely that many do not know exactly what it is or why it is so important for disease diagnosis. This technique allows millions of copies of a specific DNA (deoxyribonucleic acid) fragment of a living organism to be obtained from a small biological sample. DNA, as you know, is where the genetic material of any species is found. These exact copies allow us to identify the DNA of that disease or pathogen that we are studying. For this reason, by analysing the results, we can obtain important information such as, in our case, whether or not there are certain viruses in the Galapagos giant tortoises. This technique is very useful because it can be utilised to perform other analyses, such as paternity testing, forensic analysis, and detection of illegal wildlife trafficking, etc.

Before the PCR was invented studying DNA was very complicated because this structure contains a large amount of genetic information and detecting or isolating specific genes was a challenge. In fact, a decade before its discovery, gene cloning was the latest technology used by scientists to duplicate genetic material, but the work was laborious and very expensive. Since its creation just 32 years ago, the PCR is here to stay as a very useful technological tool, since it has managed to position itself in the field of science to this day and we have this technique to thank for the ability to detect SARS-CoV-2 in humans.

Gislayne Mendoza performing a PCR. In this step, she adds all “ingredients” into an Eppendorf tube with exact measurements previously calculated (this mixture is known as Master Mix)
Gislayne Mendoza performing a PCR. In this step, she adds all “ingredients” into an Eppendorf tube with exact measurements previously calculated (this mixture is known as Master Mix). © Rashid Cruz, CDF

Despite the fact that almost everyone on the planet has heard this word in the last two years, or even recently undergone a PCR, it is very likely that many do not know exactly what it is or why it is so important for disease diagnosis.

Carrying out a PCR is very similar to following a kitchen recipe, we need ‘ingredients’, measurements, exact times and ideal temperatures to obtain our result. Among the ingredients is the ‘template’ DNA fragment that will give us the genetic information that we need to ‘copy’. Here we can remember what we were taught in biology classes at school about this ladder-shaped molecule turned in helix format, which is part of all living beings on planet Earth. In addition, we need primers or oligonucleotides, which are short DNA sequences specifically designed to help us find the region we want to study, and starting point to generate new DNA copies. To complete our list, we need a few important ingredients to extract and copy our DNA fragment, such as the buffer solution, dNTPs, Taq Polymerase enzyme and Magnesium ions (Wages, 2005).

Once we have all our ingredients, we calculate the appropriate proportions, depending on the number of samples we want to analyse. Something similar to the quantities that we put when we prepare a cake and measure the ingredients according to the number of people. When we get our exact mix of ingredients, we dispense it into small tubes and take them to our ‘oven’, known as a thermocycler. This is a device that raises and lowers the temperature according to the programming indicated. This last step is essential to obtain the result of our long-awaited ‘cake’, since we need several cycles with ideal temperatures for the DNA chain to open, new fragments to be synthesized in each cycle and finally, those chains to close.

Gislayne Mendoza placing the sample in the thermocycler (the ‘oven’)
Gislayne Mendoza placing the sample in the thermocycler (our ‘oven’). © Rashid Cruz, CDF
Floreana giant tortoise

The return of the Floreana giant tortoise

The remarkable story of the Floreana giant tortoise, a species that the world thought was lost forever.

Read more

To visualise the DNA fragments that interest us, another technique with a galactic name, called Electrophoresis is used. This is a laboratory technique that is responsible for separating DNA molecules with the help of an electric current. For this procedure, we need a device that contains a negative charge at one end and a positive charge at the other. Therefore, when inserting DNA molecules (which have a negative charge), they will migrate to the positive side, allowing DNA fragments to be detected in the form of bands, using a contrast with ultraviolet light (Michov, 2020). In this way, we can determine if the samples we are studying are positive for a certain virus, thanks to the presence of these bands, as shown in the following image:

Agarose gel showing bands compatible with positive results of adenovirus from Galapagos tortoises at the Charles Darwin Research Station Fisher Laboratory.
Agarose gel showing bands compatible with positive results of adenovirus from Galapagos tortoises at the Charles Darwin Research Station Fisher Laboratory. © Gislayne Mendoza Alcivar, CDF

Within the GTMEP, the use of PCR has allowed us to describe four new viruses in giant tortoises: two adenoviruses and two herpesviruses. Although everything seems to indicate that these viruses are endemic to the islands, that is, they have evolved with their host (giant tortoises) over time, it is also suggested that these viruses could cause disease and leave long-term illness, especially in animals suffering from immunosuppression or stress from captivity or transport.

Ainoa Nieto providing instructions to Anne Guezou and Freddy Cabrera. An adult tortoise can be seen next to Ainoa, apparently, also paying attention to the instructions. Española Island
Ainoa Nieto providing instructions to Anne Guezou and Freddy Cabrera. An adult tortoise can be seen next to Ainoa, apparently, also paying attention to the instructions. Española Island. © Juan Manual García, CDF

The use of techniques such as PCR can help us better understand these risks. We can only conserve what we know, so it is important to continue this study in all species of Galapagos tortoise to catalogue the infectious agents that are present, and support the creation of prevention, control and surveillance plans for these diseases, so we can preserve these centuries-old species that are at risk. Thanks to my participation in this project, I have had the opportunity to gain theoretical and practical experience, both with new laboratory techniques to analyse the health of tortoises, working in the field, and supporting different investigations within the programme. In the laboratory, just like in the kitchen, we do not always get perfect results. With a lot of patience, I have been able to experiment and try different ‘ingredients’ to perfect and solve every detail, until I get ‘the perfect cake’.

Gislayne Mendoza taking morphometric measurements of a giant tortoise.
Gislayne Mendoza taking morphometric measurements of a giant tortoise. © Karina Ramón, CDF

This programme is a multi-institutional collaboration among the Charles Darwin Foundation, the Max Planck Institute of Ornithology, the Galapagos National Park, Saint Louis Zoo Institute for Conservation Medicine, the Houston Zoo and Galapagos Conservation Trust.

This blog was originally posted on the Charles Darwin Foundation website and has been edited slightly for our website. 

How you can help giant tortoises

Donate or adopt a giant tortoise and help us protect the Galapagos Islands for many tortoise-lifetimes to come.

Related articles

14th May 2024
Plastic pollution Women in science

Researching plastic and chemical pollution in Galapagos

Georgie Savage, PhD student at the University of Exeter, introduces us to her work on plastic pollution and shares stories from her recent research trip with GCT to Galapagos.
Read more
Marine iguana expelling salt from its nostrils
2nd May 2024
Technology Women in science

How can we use technology to save the Galapagos marine iguana?

Andrea Varela of the Iguanas From Above project shares her experience studying the iconic marine iguana using the latest technologies.
Read more
19th Apr 2024
Research Women in science

Studying health in birds: An interview with Gislayne Mendoza Alcívar

Gislayne Mendoza Alcívar, laboratory technician at the Charles Darwin Foundation, shares her experience studying the health of some of the Islands' most endangered birds.
Read more
Diana Pazmiño and the Gills Club (Chicas con Agallas)
26th Feb 2024
Education and outreach Women in science

Gills Club: Empowering young women in Galapagos

Gills Club (Chicas con Agallas) aims to engage more girls in shark science in Galapagos, and is supported by GCT as part of our education and outreach work.
Read more

Get the latest news from Galapagos

Join our mailing list to receive our monthly email newsletter, bringing you the latest news on Galapagos and our work to protect the Islands.

Hidden
Share This Page