DNA Hybridisation

DNA hybridisation A level Biology

In the context of this article, the single-stranded piece of DNA is the DNA probe, and the section of DNA or RNA is a particular allele.

For the probe to bind to the complementary sequence of the specific allele, the double-stranded DNA needs to be separated. Heating the DNA sample denatures the DNA molecules and separates the two strands by breaking the hydrogen bonds between them.

Then the separated DNA strands would be mixed with the probes, and the temperature would be lowered, allowing for hydrogen bonds to be formed (anneal) between complementary DNA strands. Although most of the DNA strands would pair up with their original complementary strand, some strands would bind to the probes. This binding process is referred to as DNA hybridisation.

DNA hybridisation Technique

DNA probes and DNA hybridisation can be used to locate a specific allele. This process involves many steps:

1. The sequence of the nucleotide base of the gene of interest needs to be determined. Sequence the DNA or refer to genetic libraries to obtain the sequence of the mutation of interest.
2. Then a fragment of DNA with complementary sequences to the particular allele we are interested in is produced.
3. Multiple copies of the DNA fragment are produced using polymerase chain reaction (PCR).
4. The DNA fragments produced must be labelled by adding radionucleotides (nucleotides containing ³²P) or fluorescent nucleotide triphosphates to the PCR mixture to create radioactive or fluorescent DNA probes, respectively.
5. We need to obtain the DNA that we want to analyse by extracting the DNA from the cells obtained from the individual of interest. The extract is then heated to denature and separate the two DNA strands from each other.
6. The DNA probes that we designed are then added to the extract, and the temperature is lowered, allowing for the complementary sequences to pair.
7. If the extracted DNA contains the mutation, our DNA probe would likely bind to it since it has complementary base sequences.
8. Any unattached DNA probe is removed from the mixture.
9. The hybridised DNA now will be either radioactive or fluorescent depending on what type of DNA probe we used.
10. We can use an X-ray film or use fluorescent microscopy to detect the presence of the hybridised DNA and confirm whether the suspected person had the mutation or not.

Fig. 1 - The hybridisation of DNA with a single-stranded DNA probe

Application of DNA hybridisation in genetic screening

Thanks to the work of countless scientists around the world, we now know the location and type of mutations that cause many hereditary diseases. DNA hybridisation and DNA probes can be used to screen individuals for specific hereditary conditions.

Multiple genetic mutations can be simultaneously investigated by fixing multiple DNA probes in an array on a glass slide and then adding the donor DNA to the array. Any complementary DNA sequences, if present, would then bind to their corresponding probe.

Cancer screening

Genetic screening can detect oncogenes (genes that increase the likelihood of developing cancer). There are two types of oncogenes, tumour suppressor genes and proto-oncogenes.

Therefore, any mutation that inactivates tumour suppressor genes or increases the activity of proto-oncogenes can result in uncontrolled cell division and lead to the development of cancer.

The presence of mutations in these proto-oncogenes can be detected by genetic screening. This would allow the individual with the mutation to make an informed decision about their lifestyle. For instance, they would be able to lower the risk of developing cancer by eating healthier and avoiding mutagenic substances such as cigarette smoke.

People can also undergo regular check-ups to detect cancer early and improve the chances of successful treatment.

Personalised medicine

Another advantage of genetic screening is personalised medicine. It allows doctors to treat patients based on their genotype. For instance, drug A may be more effective in treating a condition in one patient but not another patient with a different genotype. Another drug can then be used for the other patient that provides maximum efficacy in their treatment.

Knowing the patients’ genotypes allows doctors and pharmacists to prescribe the exact dose of a specific medicine to produce the desired outcome on the patients’ treatment plan. This would save money and prevent any unnecessary harm and side effects that some medications may have.

Application of DNA hybridisation in genetic counselling

Genetic counselling is a special form of social work. It provides advice and information to people, helping them make personal decisions about themselves and their children. Based on family history of hereditary conditions and genetic screening, genetic counselling can inform couples about their children’s chances of having a disease.

Here is some information to refresh your memory; alleles are usually either dominant or recessive. Suppose an individual is heterozygous for the dominant allele (i.e. has the dominant allele on one chromosome and the recessive allele on the other chromosome). In that case, they will show the dominant phenotype despite carrying the recessive allele. Only if an individual is homozygous for the recessive allele (i.e. has the recessive allele on both chromosomes) will they have the recessive phenotype.

Fig. 2 - Normal blood cell flow is compared to sickle cell

Ethical concerns of DNA hybridisation

Despite the many advantages of genetic screening, there are some major ethical considerations around genetic counselling. Genetic counsellors face a common ethical dilemma involving genetic screening on pregnant patients. We mentioned earlier that genetic screening could determine whether one or both parents are carriers of a medical problem. Other tests can determine whether the fetus has genetic mutations that will result in birth deformities, mental disability, or physical impediments. Counsellors may find it ethical challenging if they know that their pregnant patients may decide to terminate the pregnancy over carrying a fetus to term.

Inappropriate testing is another ethical challenge that surfaced when genetic testing became increasingly prevalent. This refers to couples who seek genetic counselling and testing in the hopes of having a baby with specific characteristics that they desire. Couples can undergo testing and abort pregnancies that do not result in ideal children, or even pregnancies that result in a child of the opposite sex to the one they desired.