Please post your questions for this week in the comments section. Please ‘like’ questions posted by other learners if you are also interested in having these answered.
Thank you very much for all of your questions and comments this week, I have answered as many of them as I can below. As I said last time, there are so many questions that it just isn’t possible to answer all of them (much as I would like to!), but I have tried to answer a few more this time round.
Clarification of DNA profiling terms and procedures
Firstly, a few people asked for clarification of some of the terms and procedures involved in DNA profiling, including Mercy Musau and Goda Aleknavičiūtė. The human genome contains 3 billion bases, or letters of code, and trying to analyse all of these would be extremely time-consuming, expensive and also excessive, as we can get a lot of information about the identity of an individual by looking at a small subset of the genome. Some regions of the genome, known as genes, encode proteins that need to be made in order to build a human being, and keep that human alive. These are known as coding regions, and make up only about 1.5% of the human genome. The rest of the DNA is non-coding, some of which has no function, and some of which functions as regulatory regions controlling when genes are turned on, where they are turned on, and for how long etc.
In DNA profiling, we analyse non-coding regions called short tandem repeats (STRs), which are made up of short core DNA sequences that are repeated many times. The number of times the core sequence is repeated varies among individuals, which means these regions can be used to discriminate among different people. We examine multiple STR regions around the genome, to get more information about the person whose sample we are analysing. Each of these regions is known as a locus, and this simply defines the position of the STR region in the genome, i.e. which chromosome it is on, and where on that chromosome it is. At each locus, every individual has two copies of the STR region, because everybody has two copies of each of their chromosomes, one of which they inherited from their mother, and one from their father. The two copies of the STR region are known as alleles, and a DNA profile is just a list of allele codes found at each locus, which define the number of core repeats an individual has in their two alleles at each of the STR loci examined.
To analyse how many repeat copies an individual has at each of the STR loci, we use a technique called polymerase chain reaction (PCR), which makes lots of copies of multiple STR regions in the genome, all in a single reaction. The length of these copies depends on how many copies of the repeat sequence an individual has at each STR locus, and so we can determine their two alleles by measuring the length of the copies produced by the PCR. This is done using capillary electrophoresis (CE), which separates the DNA copies by size, and Maria Cullen had a question about this. CE is a form of gel electrophoresis, which is a general term for a technique where samples of DNA are passed through a gel matrix to separate them by size. This usually involves injecting the sample into the gel matrix and applying an electric current. The electric current causes the DNA to migrate through the gel, and the speed at which different pieces of DNA move through the matrix depends on their size – small pieces move more quickly than large pieces.
One form of gel electrophoresis uses a chemical called agarose as the gel matrix, but this has now been superseded by polyacrylamide gel electrophoresis. This is because when we run DNA fragments through agarose gel we can only resolve differences in length of about 2-4 bases, whereas using polyacrylamide gel we can resolve DNA fragments that differ in length by only one base, so this gives us better discriminatory power. CE is a form of polyacrylamide gel electrophoresis, which uses very thin glass tubes, or capillaries, pumped full of polyacrylamide. The DNA sample is injected into the glass tube at one end and an electric current applied along the tube, so the DNA fragments move through the tube and are detected at the other end. The length of time it takes the DNA fragments to get from one end of the capillary to the other is related to its length, and so this is used to determine the size of each of the fragments.
Everard Lopez asked about whether there is any verification process once a DNA profile has been recovered from a body fluid or stain.
All of the techniques used in the DNA profiling procedure are validated, i.e. tested with known samples to determine that they are working correctly. In addition, labs who do DNA profiling should be accredited, which means that they are regularly checked and inspected by accrediting bodies to make sure they are carrying out procedures correctly. In the UK for example, there are fewer than 20 labs who are accredited to generate DNA profiles for loading onto the National DNA Database, and this is to ensure that the data is as reliable as possible. In addition, labs will always use positive and negative control samples to ensure that all of the regents and techniques are working properly, and that there has not been any contamination of samples. They will also usually have DNA elimination databases, which contains the DNA profiles of all of their staff and any individuals who may come into contact with evidence samples. This means that all DNA profiling results can be checked against this database to ensure that there has not been any contamination in the lab.
Changes to DNA profiling procedures Suzanna Walker and Steve Phipps also asked questions relating to this, about changes to the current DNA profiling procedure.
Until recently, the standard DNA profile in the UK consisted of ten STR regions plus a marker to determine the biological sex of the donor of the sample, i.e. whether they have two X chromosomes or and X and Y chromosome. This was known as the SGM Plus system. In the USA the standard is to test 13 STR regions plus the biological sex marker, and this is known as the Combined DNA Index System, or CODIS. Recently, labs in the UK have moved to new systems. In England and Wales, a DNA profile now consists of 16 STR loci plus the sex marker, and in Scotland we have moved to a system with 21 STR loci and three sex markers. This gives us even greater power to distinguish between different individuals, and makes the probability of finding a match between two individuals by chance – the way that DNA results are statistically presented in court – even smaller. Forensic DNA testing will probably always involve testing STR regions, but in future we may also examine some other parts of the genome sequence, and the techniques we use to examine these and STR regions are likely to get more accurate, efficient, cheaper and more automated.
Recovering DNA from different sample types
We also had a few questions about recovering DNA from different types of samples. Cheryll Rawling asked about whether there is any information in a DNA profile about what type of tissue or body fluid DNA came from. She gave an example of DNA being transferred from one person onto another when someone sneezes or coughs. Whenever we speak or cough or sneeze etc., our DNA is transferred into the environment and onto surfaces around us. If the person who was sneezed on then alleges that the sneezer attacked them, and their skin is swabbed for DNA, the profile of the individual who sneezed may well be recovered from the swab. However, there is nothing in the DNA profile that allows us to determine whether the DNA came from a sneeze or from physical contact during an attack. There are some chemical tests that can be done that indicate a particular body fluid, e.g. blood, semen, saliva is present, but these are not conclusive and there is no test for skin cells or sweat. Even if a positive test for e.g. blood is found and then a DNA profile is recovered, there is no information in the DNA profile that definitively links the DNA profile to the blood. Often this link is assumed, and sometimes this is a reasonable inference to make, but as DNA profiling techniques become increasingly sensitive, it is going to become harder for scientists to make that assumption, as we will increasingly be picking up ‘background’ DNA profiles in samples, which have no connection to the alleged incident.
Karen Drebert asked how long evidence such as DNA, but also fingerprints and body fluids such as semen would remain viable for analysis, and correctly suggested that this would depend on the conditions in which the sample had been stored.
If a sample was at a crime scene that was outdoors, and there was a lot of rain, or very high temperatures or humidity, then samples of a biological nature such as DNA, body fluids and fingerprints could deteriorate and degrade very quickly, and this is a major challenge of managing a crime scene of this nature. However, if samples were indoors, in dry environments that were not subject to major fluctuations in temperature etc., then samples could still be analysed after months or even years.
Blanche Smith asked about recovering DNA from an open can of soft drink, or from half-eaten food, and this is sometimes possible.
To recover these samples we would probably use a moistened swab or a small piece of tape to try and recover DNA from the surface of the drink can or food, and then carry out the extraction procedure on the swab or tape, as normal. I have never tried to recover DNA from the surface of food, and the success of this would depend on a lot of factors including the amount of DNA that had been transferred onto the food, and the nature of the substance being swabbed, e.g. if it was porous or not, if it contained any chemicals that might interfere with any of the DNA profile reactions. However, recovering DNA from surfaces such as the neck of a bottle or can, or the surface of a mobile phone or back of a watch is very straightforward and often produces very clear DNA profiling results. The issue with this type of sampling is that often a mixed DNA profile is recovered, e.g. if two people have touched a drinking bottle and the bottle is swabbed, we would be likely to recover some or all of both of their DNA profiles in a single reaction, and this can be very difficult to interpret as it can be very challenging to determine which components in the profile come from which individual.
Interpretation of mixed DNA samples
This difficulty with DNA mixtures is often encountered in cases of a sexual nature, because samples will often contain biological material from an individual who has been attacked, and from their attacker. Jeanne Shields asked about how a suspect’s DNA can be identified in a case of multiple rape, and this can be very challenging because if there is DNA present from more than two individuals in a sample, it becomes extremely difficult to separate out which components of the DNA mixture came from which individuals. One way that these types of samples can be analysed is to look at STR regions that are present on the Y chromosome. If a female alleges that she has been raped by multiple males, we can analyse the sample and look only at DNA that is found on the Y chromosome – this means that we exclude any DNA of the female from the analysis. As males only have a single copy of the Y chromosome, we generally expect to see a single allele at Y STR loci from each male individual who has contributed to the mixture. We can therefore look at how many different alleles we see in a mixture at these Y loci and this can give some information on the number of different males who have contributed to the mixture, and also some information about their Y chromosome profile. However, this information is quite limited so it can still be very difficult to determine whether a specific individual contributed to a mixture, but this can be useful in excluding someone as having contributed, if they have a Y chromosome allele that is not present in the mixture.
Genetic chimeras and transfusions
A few people also asked about genetic chimeras, and whether blood or other transfusions would affect the outcome of a DNA profiling test. Laura Quinoñes Urquiza, Hedley Quintana, Kenneth Ransom, Julie Webb-Pullman, Ross Thompson, Anna Dreisen, Juan Carols Alvarex, Vaughan Hnecher, A El Sokary and Stuart Gibbon all contributed questions or discussion points about this. There are two different but related issues here, one about genetic chimeras/mosaics, and one about the effect of transfusions.
Individuals can be chimeric if they have different genomes in different parts of their bodies, and these are quite rare. Individuals can also be mosaics if there are genetic mutations present in some of their cells, and this is very common – we are all likely to be genetic mosaics to some extent. Depending on whether an individual was a chimera or a mosaic, and depending on which tissues in their body had different genetic sequences, this might affect the outcome of a DNA profile. For example, if someone had a different genetic complement in their blood, saliva or semen than they did in other parts of their body, this might affect a police investigation. This was what happened in the case of Andrei Chikatilo, which Laura Quinoñes Urquiza, Hedley Quintana and Kenneth Ransom all mentioned. You can read a bit more about chimeras and mosaics and how they arise in this article.
If someone has a blood transfusion, the DNA of the person who donated the blood can persist in the recipient’s body for a while, under some circumstances. When you donate blood, it is separated into its different components, including the plasma, platelets, red blood cells (erythrocytes) and white blood cells (leukocytes). A blood transfusion is primarily made up of red blood cells, which don’t have any DNA in them, and so the majority of donated cells will not transfer any DNA, as A El Sokary correctly pointed out. However, if a patient requires a transfusion of whole blood, they will also get white blood cells, which contain the donor’s DNA. Some scientists also think that there will always be transfer of some white blood cells, so some DNA will always be transferred. Depending on how many white blood cells are transferred, the donor’s DNA might persist in the recipient’s blood for a few days and so if a blood sample was to be taken of someone who had a blood transfusion and a DNA profiling test carried out, the result might show the DNA profile of the donor, or more likely a mixture of the donor and the recipient. However, blood cells are regenerated in the body regularly, and so this effect would be unlikely to last very long, so would not have a big impact on DNA profiling tests. You can read a bit more about the effects of blood transfusion on DNA profiling in this article.
A different type of transfusion that could have a much greater impact on DNA testing is bone marrow transfusion, as this permanently changes the DNA content of some of the cells in the recipient’s body. Bone marrow transplants are used to treat certain diseases, including some cancers of the blood and bone marrow such as leukaemia. To undergo this treatment, the patient’s own bone marrow and blood cells are destroyed, usually using chemotherapy, before putting in bone marrow from the donor. The donated bone marrow cells have the DNA of the donor in them, and because bone marrow contains stem cells from which blood is made, the recipient then goes on to produce blood that contains only the donor’s DNA, permanently. This would mean that if a blood sample were taken from the recipient, it would produce a DNA profile matching the donor, and a different DNA profile than would be produced from another tissue or sample type in the recipient’s body, e.g. a cheek swab. You can read a bit more about the effects of bone marrow transplant and a sexual assault case where it arose in this article It is now sometimes the case that when bone marrow transplants are carried out, not all of the recipient’s blood and bone marrow cells are destroyed before the donor cells are transplanted, so those individuals would give samples that produced DNA profiles that were a mixture of the donor and recipient profiles.
There were also a couple of questions and comments from Sheena Middleton, Linden Brooks and Patricia Noone, about people who have been wrongly convicted as a result of unreliable DNA evidence, and those who have been exonerated as a result of DNA testing carried out on evidence that could not be tested at the time. There are many examples of old cases that have been reviewed and solved as a result of new DNA testing being carried out on old samples that were not tested during the initial investigation of a crime, either because it occurred prior to the widespread use of DNA testing, or because techniques were not sensitive enough at the time to generate a DNA profile from a small amount of sample. Often, the individual who committed a crime is caught many years later as a result of a chance encounter with the police that results in a DNA profile being taken and uploaded onto the National DNA Database, leading to a match to a profile recovered from evidence in an old case. An interesting case of this type has been reported in the UK media this week, which you can read more about in this article. There are also many examples of individuals being exonerated of crimes they have been convicted of when DNA testing is applied to old evidence samples and shows that they could not have been the source of the sample. A big part of this work is done by innocence projects, originally in the USA but now also in the UK. In the UK many innocence projects are based at universities and involve students working on real cases; you can read some more about this here.
Finally, there were a few questions and comments about differentiating identical twins using DNA testing, from Atinga Ba-Etilayoo, Rita Donà, Stuart Gibbon, Harry Nichol and Jonathan Withers. I wrote a bit about this in my answers to the ‘Ask Penny’ section in Week 1, so you might find it useful to go back and have a look at that. John Stocker also asked about sharing DNA profile records around the world, and there is some information about this in the Week 1 answers as well.
Thank you for all of your excellent comments and questions this week. I hope you are all still enjoying the course, I’m sorry again that it is not possible to answer everything, but I hope you will find this information useful as you get started on the Week 4 material.