Activity 11.1 Determining Identity based on Mitochondrial DNA Sequence Compariso

Question

Activity 11.1 Determining Identity based on Mitochondrial DNA Sequence Comparison In this activity you will be provided with a reference source of mtDNA from a mother who lost two siblings during wartimes. You will be provided with her mtDNA sequence for reference purposes and mtDNA sequences extracted from two different burial sites where her sons were known to have fought during wartimes. Your objective is to (1) determine all three mtDNA sequences and (2) perform a mtDNA sequence comparison to determine if these skeletal remains are her sons Materials: One set of known maternal mtDNA sequence Two other unknown sets of mtDNA sequences Protocol The instructor will provide you with three different colored sets of mtDNA bases that need to be assembled using the concept of Contigs performed in a previous activity. (note: in this activity a Contig must consist of a minimum four (4) base overlap region) 1. 2. One set represents the mother 's known reference mtDNA sequence. Two other mtDNA sequences represent skeletal remains exhumed from two different burial sites believed to be the remains of the mother's missing sons. 3. Using the data table below write out the sequence of the mother's DNA followed by the mtDNA sequences from the two different burial sites. 4.

Explanation / Answer

Hi,

1) In the activity mentioned above we can write same referance sequence available to two sons because mitochondrial sequence will be same in both siblings and it can be transfered only from mother during zygote formation. This is also termed as mitochondrial inheritance.

2) In addition to energy production, mitochondria play a role in several other cellular activities. For example, mitochondria help regulate the self-destruction of cells (apoptosis). They are also necessary for the production of substances such as cholesterol and heme (a component of hemoglobin, the molecule that carries oxygen in the blood).

Mitochondrial DNA contains 37 genes, all of which are essential for normal mitochondrial function. Thirteen of these genes provide instructions for making enzymes involved in oxidative phosphorylation. The remaining genes provide instructions for making molecules called transfer RNAs (tRNAs) and ribosomal RNAs (rRNAs), which are chemical cousins of DNA. These types of RNA help assemble protein building blocks (amino acids) into functioning proteins

13 of its 37 genes are involved in the process known as oxidative phosphorylation. This is the metabolic pathway that produces adenosine triphosphate (ATP), the main energy source of the cell. The remaining 24 genes are involved in the creation of transfer RNA (tRNA) and ribosomal RNA (rRNA) which help to turn amino acids into proteins.

3)

Mitochondria are inherited only in the maternal ova and not in sperm. Therefore, a pattern of inheritance associated with alterations in mitochondrial DNA gives a pattern of the condition affecting males and females, but always being maternally inherited. An affected male does not pass on his mitochondria to his children, so all his children will be unaffected.  This is called mitochondrial (sometimes matrilineal) inheritance.

It is known that there are some mitochondrial conditions associated with incorrectly functioning mitochondria where the mutation is not in the mitochondrial DNA, but in genes in the cell nucleus that control functions in the mitochondria. It is therefore possible for a mitochondrial condition not to show mitochondrial inheritance but to show Mendelian inheritance.

Mitochondrial DNA VS Nuclear DNA

Mitochondrial DNA:

It is found in the mitochondria of a cell. Mitochondrial DNA has only one chromosome which is comparatively shorter and they do not code for all the proteins but only specific proteins. This DNA make its own tRNA and even rRNA. It replicates separately from the nuclear DNA and is passed on to the progeny from mother only. That being the reason for which it does not undergo recombination. It is even read with a different genetic code from the nuclear DNA itself.

Nuclear DNA:

Unlike mitochondrial DNA, nuclear DNA is found in every cell of the body. It consists of 46 chromosomes which are inherited from mother and father, both. The are comparatively longer with centromeres and telomers. DNA is composed of the adenine, thymine, guanine and cytosine which are the main nucleotides. DNA consists of a double helix chain, containing histones. This double helix strand is separated during division by various enzymes. The nucleotides are paired along the whole length of DNA, any change in the sequence of these nucleotides could prove very disturbing and alarming.

Differences:

4) Each sibling gets the same distribution of parental DNA in terms of percentages. That is, each sibling gets half of their DNA from the mother and half from the father*.

However, each sibling gets different parts of the two parents' DNA. It always adds up to 50:50, but if you have a gene where the father has the two versions A and a and the mother has the two versions B and b, you could have siblings that end up with AB, Ab, aB, or ab. Each sibling has one version of the gene from their mother and one from their father, but the combinations are different. The same's true for every gene, because how the chromosome sections separate out as the egg and sperm cells are formed is essentially random. This is why you end up with siblings who are similar, but not identical - they're made from the same starting material (the parents' genes), but in different combinations.

Male offspring actually have slightly more DNA from their mother than from their father, because the X chromosome they get from their mother is bigger than the Y chromosome they get from their father.

5)

Nuclear DNA is analyzed in evidence containing blood, semen, saliva, body tissues, and hair follicles. DNA from the mitochondria, however, is usually analyzed in evidence containing hair fragments, bones, and teeth. Mitochondrial DNA analysis is typically performed in cases where there is an insufficient amount of sample, the nDNA is uninformative, or if supplemental information is necessary.

Unlike nDNA, where one copy of a chromosome comes from the father and the other from the mother, mtDNA is exclusively inherited from the maternal side. Therefore, the maternal mtDNA should be the same as her offspring. This can be helpful in cases where it is not possible to obtain a sample from the suspect but it is possible to obtain a sample from one of the suspect's biologically related family members. By doing so, the suspect can be excluded as the culprit of a crime if the results indicate that the relevant family member's mtDNA does not match the mtDNA fingerprint from the sample.

Mitochondrial DNA can be informative in a different way than nDNA. Less than 10% of the mitochondrial genome is noncoding and localized in a region called the D-loop. In this region, there are sequence variations that are inherited that can be used for forensic purposes. These regions, called hypervariable regions, are broken down into two sections: HV1 and HV2. It is within these regions that inherited sequence variations can be identified.

One of the main reasons mtDNA analysis can be helpful to forensic scientists is that in some tissues, mitochondrial DNA is in excess compared to nDNA. As nDNA exists in chromosomes and there are only two copies of each chromosome (one inherited maternally, the other paternally) per cell, the nDNA copy number is much smaller. The mitochondrial genome can have a copy number of 2–10 per organelle and in some cases the number of organelles can reach the hundreds. For example, in muscle tissue, where the demand for energy is highest, there are a larger number of copies of the mitochondrial genome. Analysis of mtDNA, therefore, can be particularly helpful in forensic cases where sample integrity or size is compromised or when confirmation is needed.

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