cellular and molecular

Question 1

DNA is made up of 4 deoxyribonucleotides containing the purine bases _______ and ________, and the pyrimidine bases _________ and ________.

The bond between the bases is a ________ bond.

Answer 1

adenine and guanine
cytosine and thiamine

Adenine binds to thymine and cytosine binds to guanine.

hydrogen

Question 2

2. The central dogma of molecular biology is that DNA can be copied to DNA and to messenger RNA (process is called _________), and that proteins can be synthesized using the information in mRNA as a template (called _______), but that the information can’t be transferred back from protein to nucleic acid, or from RNA to DNA.

Answer 2

transcription, translation

Question 3

3. How is the structure of RNA different from DNA?

Answer 3

• Thymine is replaced with uracil
• The base pairs are linked by ribose rather than 2’ deoxyribose
• RNA is usually single stranded
• RNA is much more susceptible than DNA to degradation by nucleases

Question 4

4. What are/were the benefits of sequencing the genome?

Answer 4

• The expectation that knowledge of the sequence of position of all genes would produce tangible improvements in medical care
• Tools could be developed for storing and analyzing the large amount of information produced
• The work would produce a biotechnology industry to stimulate the development of new medical applications from the data

Question 5

5. A Boxer was chosen for the canine genome sequencing project because this breed was described as having the lowest rate of _________, in other words, the lowest rate of variation in sequence, when compared to other breeds.

Answer 5

heterozygosity

Question 6

6. The feline genome has also been sequenced. What is of particular interest?

Answer 6

-The large numbers of endogenous retrovirus-like sequences identified, accounting for about 4% of the feline genome.

Question 7

7. Only approximately __% of the genome codes for mRNA, rRNA, tRNA, small nuclear RNA, and small nucleolar RNA.

Answer 7

10

Question 8

8. Transcription is mediated by a DNA-directed what? Where in the cell does it occur?

Answer 8

• RNA polymerase

- Primarily in the nucleus, but also in the mitochondria

Question 9

9. The length of genes is often many times greater than that of the transcribed mRNA molecule due to the presence of _______ which are noncoding nucleic acids, that separate the exons, or the coding sequence. The base triplets of three nucleotides are termed codons and code for individual amino acids.

Answer 9

introns

Question 10

10. Translation of mRNA molecules to a polypeptide occurs in the _________.

Answer 10

ribosomes

Question 11

The presence of intronic sequence permits alternative splicing of the exons and thus variation in the sequence, which is translated to protein from a single gene and is called a ____ ______. An important example is the increased expression of a splice variant in breast cancer causing a reduced survival time due to shorter time to tumor mets.

Answer 11

splice varient

Question 12

12. The majority of genomic DNA present within mammalian cells is not _______, with less than 2% of the haploid human genome coding for genes.

Answer 12

transcribed

Question 13

13. Areas of noncoding elements, which are highly conserved between mammalian species, are often associated with genes that code for ____________________

Answer 13

regulation of development.

Question 14

14. The one way flow of genetic information is not always the case. What is the enzyme that can generate a DNA sequence from and RNA template? What is the best described reverse transcriptase in mammalian cells and why is it thought to be important?

Answer 14

• Reverse transcriptase
• Telomerase, implicated in aging and oncogenesis b/c it is responsible for preventing the shortening of the telomere length and its expression is a near universal marker of malignancy

Question 15

15. Segments of DNA sequence that are transcribed into mRNA are termed _______. Its position and the start of the gene are identified by a sequence upstream termed the _______.

Answer 15

transcription units

promoter

Question 16

16. Genes such as ribosomal proteins and histones have a common function between different cells and thus are constitutively expressed between different cell types and are called ______ genes.

Answer 16

housekeeper

Question 17

17. What is epigenetics? What are some of the mechanisms by which it occurs? Give an example in dogs.

Answer 17

• Methods other than DNA sequence that control differential expression in different cell types within an individual
• Via DNA methylation and histone acetylation
• Hypomethylation is associated with transcriptional activity and has been implicated in the neoplastic transformation of cells and has been identified in canine neoplasia such as lymphoma

Question 18

18. What are mutations called that don’t affect the fitness of an individual?

Answer 18

-Neutral mutations

Question 19

19. What is a haplotype?

Answer 19

-When a set of alleles that are closely linked at a particular locus and they are inherited together, each different set is termed a haplotype

Question 20

20. What is the most common type of mutation? What are other examples.

Answer 20

-Most common is a single nucleotide polymorphism, or a point mutation
If it changes a protein sequence or length it is called a nonsynonymous mutation
-A missense mutation results in a change to an amino acid codon, which may alter the protein structure
-A nonsense mutation is when the mutation may replace the normal amino acid codon with a stop codon leading to termination of the protein sequence
-A synonymous mutation changes the sequence but not the amino acid
-The deletion or addition of a single or multiple base pair sequence will change the frame in which the sequence is read by RNA polymerase and is termed a frame shift mutation

Question 21

21. What is genetic linkage?

Answer 21

-The tendency to inherit together two or more alleles at different loci on the same chromosome more frequently than would be expected by independent assortment.

Question 22

22. What is linkage disequilibrium?

Answer 22

-The association of alleles at different loci, but not necessarily on the same chromosome, at greater frequency than would be expected by random chance.

Question 23

23. What are some ways of detecting mutations?

Answer 23

• Use of restriction nucleases which are bacterial enzymes that cut the DNA double helix at restriction sites and identify mutations resulting in loss or gain of a restriction site.
• DNA sequencing

Question 24

24. What is the purpose of a gene knockout experiment?

Answer 24

-They involve the insertion of a mutated (nonfunctioning) gene into a vector, which is then inserted into embryonic stem cells, leading to offspring with the gene knockout. This is the gold standard for determination of the effect of the loss of a single or both copies of a gene, or the mutation of a gene on an organism.

Question 25

25. What enzymes are responsible for the posttranslational modification of a protein by the addition of a phosphate group? What does this phosphorylation do functionally?

Answer 25

- Kinases - Causes the protein to interact with other proteins recognizing the phosphorylated domain and is commonly associated with cell signaling

Question 26

26. What are examples of tests utilizing proteomics in veterinary medicine?

Answer 26

-Western blot, immunohistochemical staining, ELISA, mass spectrometer

Question 27

27. What is pharmacogenomics?

Answer 27

-The study of the interaction of genetic variation with response to a drug, be that efficacy or toxicity

Question 28

1.) What is a biomarker, according to the NIH?

Answer 28

a. Characteristic that is objectively measured and evaluated as an indicator of normal biologic processes, pathogenic processes, or pharmacologic responses to therapeutic intervention

Question 29

2.) Where is the greatest proportion of cartilage in the body (which makes specific OA biomarker development difficult)?

Answer 29

a. Spine and respiratory system

Question 30

3.) What does the Dimethylmethylene Blue (DMMB) assay measure and how does it work?

Answer 30

a. Measures intact GAG chains via colorimetric assay to the insoluble precipitate formed by DMMB and intact GAG chains

Question 31

4.) What are the general targets of the aggrecan assays (there are 3 main sequences the assays are made toward)?

Answer 31

a. Repeating disaccharide sequence that can be sulfated at 4- or 6- on the chondrointin chains (7D4 and 3B3 +/- CS846) b. Keratan sulfate (5D4) c. N-terminus of cleaved aggrecan (BC-3, BC-14, OA-1)

Question 32

5.) What is removed from the Type II collagen pre-propeptide to allow it to form the triple helix?

Answer 32

a. C- and N-terminal globular domains

Question 33

6.) What does MMP stand for and how did they get that name?

Answer 33

a. Matrix metalloproteinases- have a metal ion, usually zinc, present at the active site

Question 34

7.) What is the result of collagen proteolysis and why is that significant for biomarkers?

Answer 34

a. Triple helix cleavage results in a ¾ fragment and ¼ fragment, which exposes the N- and C- terminal epitopes for detection (target of most assays)

Question 35

8.) What does Cartilage Oligomeric Matrix Protein do?

Answer 35

a. Binds Tpye I or Type II cartilage and stabilizes the network b. Facilitates fibrillogenesis

Question 36

9.) How are Acute Phase Proteins described (2 specific variables)?

Answer 36

a. Positive or negative based on response to insult | b. Major, moderate or minor based on magnitude of response

Question 37

Name 3 main acute phase proteins.

Answer 37

a. Albumin, Alpha-1 acid Glycoprotein, C-reactive protein, Ceruloplasmin, Haptoglobin, Serum amyloid A, fibrinogen, transferrin

Question 38

What are the stimulants of TNF-α production and where is it synthesized?

Answer 38

a. Lipopolysaccharide and IL-1 | b. Macrophages

Question 39

13.) What triggers IL-6 production from what source?

Answer 39

a. Pathogen-Associated Molecular Patterns (PAMPs - component of specific microbes) trigger IL-6 production via toll-like receptors b. Sythesized by white blood cells, mainly macrophages and T-lymphocytes, and somewhat by WAT (white adipose tissue)

Question 40

14.) Name the three primary adipokines associated with obesity related insulin resistance.

Answer 40

a. TNF-α, IL-6, PAI-1

Question 41

Obesity is characterized by a state of chronic__________, with increasing levels of inflammatory markers such as 1) 2) 3)

Answer 41

low grade systemic inflammation 1) C-reactive protein 2) IL-6 3) TNF-α.

Question 42

16.) What adipokines have been detected in the synovial fluid of human osteoarthritis patients?

Answer 42

a. Leptin | b. Adiponectin

Question 43

1.) What are the three categories of stem cells that have been identified?

Answer 43

a. Embryonic b. Fetal or perinatal c. Adult (mesenchymal/stromal, hematopoietic, and neural)

Question 44

2.) Define stem cells

Answer 44

a. Undifferentiated cells that possess the ability to divide for indefinite or prolonged periods in culture and may give rise to highly specialized cells characteristic of each embryonic germ layer b. Undifferentiated cells that possess the ability to divide for indefinite or prolonged periods in culture and may give rise to highly specialized cells characteristic of each embryonic germ layer c. The inner cell mass of preimplantation blastocysts

Question 45

3.) How are hematopoetic stem cells typically used?

Answer 45

a. Implanted into leukemia patient after immunodepletion via total body irradiation

Question 46

4.) What is so important about umbilical cord blood-derived stem cells?

Answer 46

a. More effectively repopulate the hematopoietic cell system and with less immunorejection than other stem cell sources

Question 47

5.) What do mesenchymal/stromal stem cells do besides replace a cell population?

Answer 47

a. Secrete trophic factors that affect surrounding cells | b. Secrete cytokines and chemokines to modulate immune and inflammatory reactions

Question 48

6.) Induced pleuripotent stem cells (iPS cells) are produced by…

Answer 48

a. Reprogramming of adult or embryonic somatic cells via transduction of a specific set of transcription factor genes

Question 49

7.) Why are iPS cells so important?

Answer 49

a. Mitigates ethical concerns around embryonic stem cells b. Uses autogenic transplantation so decreases immunorejection potential c. Can still be induced to differentiate into multiple lineages

Question 50

8.) What is the largest concern regarding the use of stem cells successfully?

Answer 50

a. Concern for tumor formation

Question 51

9.) What factors influence stem cell survival?

Answer 51

a. Cell delivery b. Biomaterials scaffold c. Growth factors d. Mechanical forces