Bio Exam 4

Compare DNA in prokaryotes and eukaryotes. What's the significance here?

Prokaryotes: 1 circular DNA molecule that is relatively small and attached to the cell membrane, not contained in any nucleons and no histones Eukaryotes: many large linear DNA molecules contained within a nucleus with histones. The significance here is that eukaryotes have a bigger task. They must replicate and distribute more and larger DNA molecules.

When does recombination happen, and what is also present for this to occur?

Recombination happens in prophase 1 after DNA replication, and enzymes are present in meiosis to break and rejoin dsDNA

Making a library with cDNA (complementary DNA). What's different about cDNA? When would we do this, specifically?

Use reverse transcriptase to generate DNA from mRNA. Chew up (get rid of) mRNA Allow cDNA to duplicate. The unique thing about cDNA is that it only has exons. No introns because mRNA got rid of the introns. This makes cDNA more manageable and less complex. We can do the same thing we did with gDNA where we cut and transform into plasmids and into host cells to make cDNA library. We would do this specially when we want to analyze TRANSCRIBED genes.

Lm* which is more likely to produce functional proteins in a bacterium. cDNA or gDNA? Why?

cDNA. cDNA (made from reverse transcriptase of RNA) has no introns and is less structurally comped. Bacteria don't have the enzymes required to splice out introns.

How many centromeres does a single chromatid have? How many centromeres does an X shaped chromosome have?

1 for each. 1 chromosome has 1 centromere no matter if that is a single chromatid or a pair of sister chromatids.

Consider two co-dominant alleles. How many total alleles of one gene would there be in a diploid organism with a population of 100?

100 individual: 2 copies of each chromosome. 200 total alleles.

What do you need for Sanger DNA sequencing?

A DNA template of unknown sequence. A DNA primer to initiate hybridization of this unknown sequence. DNA polymerase to add NTPs Normal dNTPs A small amount of special ddNTPs Gel electrophoresis and camera

What is a chromosome? A chromatid? How many of each before S? After S?

A chromosome is a DNA molecule. A chromatid is one double stranded DNA molecule. Before S, there is 1 chromosome and 1 chromatid. After S, there is still considered to be 1 chromosome but now there are two "sister" chromatids. Each of these sister chromatids are identical and each have double stranded DNA molecules.

What is a back cross?

A cross to a homozygous parent.

What phenomenon explains the tortoise shell cat? What is the physical correlate of this?

A female who received two X chromosomes. One from mother and one from father. Instead of codoninace, one X chromosome is on and the other is off IN A GIVEN CELL. In the zygote, both Xs are on but in a few hundred cells one X is inactivated. X inactivation is random but stable. A "Barr body" is the physical correlate. Basically a black body that binds and inactivates one of the X genes on the chromosome.

Meisosis. What is it? What cells undergo Meisosis? What are the stages?

A germ cell goes from diploid to haploid to make gametes. A reshuffling of chromosomes and alleles happens. It consists of 2 consecutive rounds of cell division following one round of DNA replication. In Meisosis 1, homologous chromosomes separate. In Meiosis 2, sister chromatids separate (in this way, it is said that meiosis 2 most resembles Mitosis). The result is 4 haploid meiotic products.

What is important to remember about DNA when it comes to its weight, especially in the context of adding heat?

A heated DNA strand, whether it's a circular plasmid or a DNA fragment, will denature in heat and the two strands will come apart. That's why, in problem 13-7, a plasmid with a weight if 5 million gives you 2 2.5 million strands when it's heated. Heat denatures the hydrogen bonds that hold the strands together.

What can be said about frequency of recombination (the word for non parental gamete formation) in terms of how far apart two genes are. What is the equation for recombination frequency?

A recombinant forms when a crossover even occurs between two genes. This is more likely to happen if the genes are far apart. Recombination frequency is equal to the #recombinant gametes over the total # of gametes x 100%.

What is a testcross? What does it tell us?

A testcross is a cross with a homozygous recessive trait (ex: bb) This is used if we want to determine if an organism exhibiting the dominant trait is homozygous or heterozygous. If the organism with the dominant trait is homozygous, all the offspring will exhibit this trait. If the organism with the dominant trait is heterozygous, half the offspring will exhibit this trait and the other half will exhibit the recessive trait.

Replication, transcription, translation, and proper function. What can human genes accomplish in a bacterium?

All but proper function. If there's an origin of replication, human DNA can be replicated as part of a plasmid. It can also be transcribed and potentially even translated. The issue arises mainly with introns - and how the translation of introns isn't conducive to making functional product by the very nature of what introns are.

In terms of p, q, and p squared, q squared, and 2pq, what are we talking about if we're talking about allele frequency? What about individual's phenotypes?

Allele frequncy, p or q. Phenotypes equal the squared numbers or 2(pq)

What is an easy way to determine if a cell is in Interphase or in Mitosis/Meiosis?

Are chromosomes visible, or is it spaghetti like (interphase).

What is important to remember about bacterial DNA?

Bacterial DNA does not have introns. It is equivalent to cDNA in this way. We looked at an example of transforming plasmids with bacterial insulin genes to select for and produce large amounts of insulin.

Why do we say that RF underestimates crossover events? How does this mess with our estimation of distance between genes?

Because of multiple crossover events. If two genes are very far apart, relatively, two recombination events can occur that essentially undo each other so no recombinant is detected but multiple crossover events occurred. This messes with our estimation of distance because we'd be more likely to assume to genes are closer together because no recombinant was detected, but really they're far apart and two crossover events occurred.

What is a aneuploidy?

Chromosome number not a multiple of N

What are the three doubles for a diploid organism?

Double stranded DNA makes up a chromatid Two sister chromatids in a chromosome. Two homologous chromosomes in a diploid organism.

When discussing mitosis, you must be talking about a _____ cell

Eukaryotic. No mitosis in prokaryotes.

True or false: C is a unit less variable.

False. C is the DNA content of a haploid cell. It's typically measured in grams or picograms.

What is pedigree analysis, and what is important to remember about pedigree analysis questions?

Family tree. Often dealing with grey area terminology.

What is the implication of the fact that RF underestimates crossover events?

For genes that are far apart, we lose the ability to detect crossovers. This is just another way of saying our data isn't reliable.

What can be said about the number of chromosomes/chromatids a cell must have ti successfully undergo mitosis and meiosis?

For mitosis, since this is all about sister chromatids separating, you need an even amount of sister chromatids that can be divided equally. For Meisosis, since Meiosis 1 is all about homologs separating, you need an even amount of chromosomes that can be separated evenly.

If you're in a situation where you see a population is not in genetic equilibrium, what is the first thing you should think of?

For some reason, one of the phenotypes is being selected for.

Eukaryotic Cell Cycle. What phase would a radioactive T be incorporated?

G1: normal cell activity. Preparing for DNA replication. S: DNA replication/synthesis G2: normal cell activity, preparing for mitosis M: Mitosis. Where 1 cell becomes 2. Radioactive T only incorporated during S phase since this is when DNA is being replicated and new base pairs are incorporated.

How can we measure recombination frequency using a testcross?

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What does the graph of distance between genes on the x axis and RF on the y axis look like in detail?

GET CLARITY HERE. BE ABLE TO EXPLAIN THIS GRAPH

What's another way to describe recombinants?

Gametes that form as a result of a crossover event in between two genes so that the gamete does not resemble either true breeding parent.

What happens when a crossover event does occur BETWEEN two genes. Use the words parental and non parental. Use the word recombinant as well.

Half the gametes have parental configuration and the other half have non parental. When we draw the crossover icon in between genes (recall this is happening between homologs not chromatids) everything below the crossover event switches sides. The 2/4 gametes that are non parental are called recombinants.

You can't use HW law if you aren't in _____

Genetic equilibrium

What is the result of a nondisjunction event in Meiosis 1?

Homologous chromosomes don't separate, so 2 gametes end up in 2 cells leaving 2 other cells without gametes.

What can be said about the number of normal gametes if ND occurs at Meisosis 1? What about meiosis 2?

If ND occurs at M1, all 4 gametes will be non normal. 2 will have one extra and 2 will have one less. If ND occurs at M2, 2 will be normal and 2 will be non normal. Of the two non normal, one will have one extra and the other will have one less.

We looked at a special case of two gene independent assortment where the genes act in the same pathway AND the same enzyme of that pathway. That is, both genomes need to be AT LEAST heterozygous if not homozygous dominant for the pathway to work. What are the odds here for the pathway working?

In this case, the only option that will work is A_ and B_. This is the common 9 option. So, the other 7/16 won't work. So you get a 9:7 success to unsuccessful ratio.

Eukaryotic DNA has two states, generally. What are they and what is the composition of DNA in these two states?

Interphase and Mitosis. In interphase, DNA is spaghetti like with no distinct structure. Polymerase can access it in this state since it is spread out. Nuclear membrane in tact. In mitosis, DNA is in sausage like chromosomes which compact together. Polymerase cannot access, and the nuclear membrane gets broken down.

Go through Meisosis in terms of what N and C are.

Interphase. N = 2, C = 2. After S phase: N= 2, C = 4 After meiosis 1: N = 1, C = 2 After meisosis 2: N = 1, C = 1

Stages of mitosis and what's going on at each.

Interphase: spaghetti like chromatic. After S, chromatin have sister chromatids. Prophase: homologous chromosomes match up, nuclear membrane breaks down Metaphase: chromosomes align in the middle and spindle fibers attach to centromeres. Anaphase: sister chromatids pulled apart. Telophase: two new nuclear membranes form. Two cells essentially ready to break away and form.

DNA Sequencing method 2: Massively parallel sequencing or Next Generation Sequencing (NGS)

Isolate DNA Generate smaller fragments Each fragmented ligated to an "adaptor" DNA fragments with adapters will hybridize to corresponding adaptor that is attached to plate (fragments sticking straight up from plate like hair standing up) DNA amplification by PCR to create lots of each fragment (still on plate) that can be detected by a microscope Sequencing now done with special, fluorescently labeled NTP. This special base also prevents another base of joining. A signal is picked up and the fleuroescent base is noted. Next, a new fluorescent base adds to the next base, and so on. Keep in mind we need a primer to start this process.

Making a library with gDNA (genomic DNA)

Isolate genomic DNA Chop up with REs Ligate into cut plasmids Transform (insert into host cell) Plate Select for trait. The point here is that each colony with a unique plasmid has a fragment of gNA. Each plasmid can be thought of as being a book in the gDNA library.

Describe what would happen if a male GERM cell underwent an nondisjunction event in Meiosis 1.

It's make so germ cell has x and Y chromosome. NDJ 1 means that homologs don't separate (remember X and Y act as homologs for all intents and purposes). So, you end up with 2 gametes out of 4 with both X and Y chromosomes and 2 gametes out of 4 with nothing. Final result would be a zygote that has 2 X (one from mother, one from father, and one Y from father)

One chromosome (with no crossing over) contains the genetic information from how many parents?

Just one. Sister chromatids are identical. The genetic information from the other parent is on the chromosomes homolog.

What is required to hold a fragment to the sticky ends of a plasmid?

Ligase. Without ligase, any sort of heat or denaturing will remove the fragment from the plasmid. P

What MUST be considered if two genes are on the same chromosome.

Linkage. This is the idea of crossovers occurring between NON-SISTER chromatids on every pair of homologs. We often drew homología chromosomes in different colors, so an easy way to think about linkage or crossovers is the sharing of colors. Sister chromatids would be the same color.

What is the likely genotype of someone with a RARE dominant autosomal condition?

Most likely heterozygous. If it's very rare, it's unlikely that an individual is homozygous dominant.

What is N? What is ploidy? What is a karyotype? What are these numbers for humans?

N is the number of TYPES of chromosomes. Ploidy is how many of each type. A karyotype is the visualization from largest to smallest of all the chromosomes in the organism. In humans, N= 23. Ploidy is 2 (2N). A ploidy of 2N is "diploid".

Life cycle of diploid animal, haploid life cycle, alternation of generations life cycle (plants)

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What is the Hardy-Weinberg equation? Define each term. What can also be said about p and q?

P squared plus 2(pq) plus q squared equals 1. P is equal to the allele frequency's of A and q is equal to the allele frequency of a. P + q is equal to one! Because these are the only two alleles available. Every individual must have at least one of these.

Define population, genotype frequency, allele frequency, and gene pool.

Population: same species in the same area that can mate with each other. Genotype frequency: fraction of population with a particular genotype Allele frequency: fraction of gene copies of a given allele Gene pool: total of alleles carried in breeding members of population in one generation.

What is important to remember about promoters, particularly in the context of cutting plasmids with REs and inserting human genes.

Promoters are (usually) directional. Often times a plasmid can be cut resulting in a straight line. If the promoter is in between two genes, only one of them might be transcribed because one of them lies in the correct direction of the promoter.

Explain this: "for genetic conditions that are x linked, you often see the condition in males"

Since females get two X chromosomes, at least one of the chromosomes will have the gene to avoid whatever condition is at hand. Since males only get 1 X, and the Y is useless in this case, half the males will have the condition.

What is the result of a nondisjunction event in Meiosis 2?

Sister chromatids don't separate, leaving one side (because it's rare) with both sister chromatids in 1 and an empty cell in the other. Other side is fine (again, because this is rare)

Compare the primary transcript with mRNA

The primary transcript refers to whatever is transcribed. All bases between the start and stop codons will be transcribed. Next, the introns are spliced out. This splicing of the primary transcript is the difference that leads to the mRNA. No introns in mRNA.

What can be said about the proportions of phenotypes in future generations in a population that's in genetic equilibrium?

The proportions of phenotypes will stay the same - especially true if the population is large enough.

What is meant by independent assortment

The random distribution of the homologous chromosomes into daughter cells during meiosis I.

What are our two option after Meiosis 1 when dealing with TWO GENES in different chromosomes. What are the possible results in Meiosis 2 from these two options? Use the words parental and non parental. How likely is each outcome?

The two genes from the same parent (blue vs red, for example) can either end up in the same daughter cell or in different daughter cells. If they end up in different, the other parents alleles ended up in different too so the result is mixed for both daughter cells. In meiosis 2 now, if the genes from one parent ended up together, the gametes will have 1 of each from one parent. In this way, all 4 gametes are "parental" If the genes from one parent ended up mixed after meiosis 1, the gametes will have 1 of each from each parent. In this way, all 4 gametes are non parental. Each outcome is equally likely, so in the long run our 8 possible gametes (4 from each type of meiosis 1 separation) will be in equal proportions.

In mitosis, in what phase do homologous chromosomes pair?

They don't. In mitosis there is no pairing of homologs. Think about it this way, the only reason homologs pair up is so that they get evenly distributed to two daughter cells. In mitosis, only sister chromatids are being separated, so there's no need for homologs to pair up together.

We looked at another special case of independent assortment with two genes where both genes incide for proteins in the same pathway but act on different enzymes. If you only encode the first enzyme correctly, you'll get a middle ground option between option 1 and 3. What genotypes will give you option 1, 2, and 3?

To get option 3 (fully successful pathway) you'll need to be A_ and B_. This is 9/16. To get option 2 (first step worked, second stop didn't work) you'll need A_ and bb. This is 3/16. To get option 1 (neither step worked). You'll need aa and B_. This is because you need the first step to work to even have the option of the second step working.

What happens when a crossover event occurs NOT between two genes. Use the word parental or non parental.

Transfer of chromosome still happens, but no transfer of these genes. So, all gametes still have parental configuration.

DNA sequencing method: Sanger DNA sequencing

Using special nucleotides dideoxyNTP in place of deoxyNTP. The difference where is that there is just an H in the 3 prime position instead of an OH. Because of this, no more nucleotides can be added AFTER this ddNTP (they can still be added to a chain due to their normal 5 prime ends - which is key). A DNA template of unknown sequence is added along with a DNA primer to intimate hybridization. DNA polymerase is needed to add normal dNTPS as well as a small amount of ddNTPs (special chain ending NTPs) These ddNTPs are dyed a corresponding color. In the long run, a ddNTP will end every sequence of different lengths, and the colors will tell you the order of base pairs.

What is co-dominance? What is an example we discussed in class?

When you see a mixture of both traits and are clearly shown. An example we discussed was blood time. A and B were codominant over i. If A and B were the genotype, both A and B sugars were added to the red blood cell. Both and A and be are completely dominant over i.

How do we determine probably of one thing and another? This is relevant when discussing two genes at a time. What common ratio does this lead to when discussing two genes?

We multiply the individual probabilities. Recall the general principles and distribution of genotypes when dealing with ONE gene. When dealing with two, the probabilities of each are multiplied. Homozygous are 1/4 and heterozygous are 2/3. This leads to the common 9:3:3:1 phenomenon. A_ B_ is 9. A_ or B_ and homozygous of the other is 3. And homozygous if each is 1. These are all out of 16.

What is directional selection?

Where one extreme is selected for, so over time the graph moves in one direction to the left or right.

What is stabilizing selection? What is an example we looked at?

Where the "middle of the road" option is selected for. Over time, the graph gets taller and narrower. An example we looked at was birth weight. The healthiest babies are average sized, and babies that are too large or small have a higher mortality rate.

What is disruptive selection?

Where the two extreme ends are selected for, and slowly two peaks form with a dip in the middle.

What are the sex chromosomes for females? For males?

XX for females. XY for makes. X and Y act like homologs in meiosis.

Why would an mRNA be longer than you would think if you estimated the length of mRNA based on the amount of codons and/or amino acids?

You have to account for the 5 prime and 3 prime UTR. These are both about 100 bases, so 200 based in total go untranslated in the mRNA when making the final peptide.

What is incomplete dominance? Also known as partial dominance.

a heterozygous phenotype is somewhere between the two homozygous phenotypes

CRISPR/Cas9

a unique technology that enables geneticists and medical researchers to edit parts of the genome by removing, adding or altering sections of the DNA sequence Very simply, CRISPR is a system that can go into the huge genome and find specific places and make small cuts and edits Cas9 protein has a guide RNA that essentially tells the protein where to go and what to cut out. That's what makes CRISPR so powerful: this little piece of guide RNA is easy to make. We can direct CRISPR to different places based on this guide RNA. After a cut, DNA can be repaired "sloppy" where mutations occur or "homology repaired" where a specific and intentional sequence is added back into the genome to correct the cut.

What is complete dominance?

heterozygous condition wherein the allele that is regarded as dominant completely masks the effect of the allele that is recessive

What is autosomal inheritance?

inheritance of genes located on non-sex chromosomes