chapter 5

Question 1

what is gene expression?

Answer 1

• explains why virtually all cells within the body contain the same genetic code but have drastically different morphology and functionality

Question 2

what is cell differentiation?

Answer 2

• describes how embryonic stem cells first differentiate into the ectoderm, mesoderm, and endoderm and then into the diverse range of specialized cells present in the mature body

Question 3

what are stem cells?

Answer 3

• relatively undifferentiated cells that can differentiate into more specialized cells and reproduce through mitosis- also present in adults

Question 4

what are somatic stem cells?

Answer 4

• come in a variety of types
  
  • hematopoietic stem cells which differentiate into various blood cells
  • intestinal stem cells which provide the basis for the constant renewal of teh cells lining the surface of the intestines
  • mesenchymal stem cells which are capable of differentiating into a wide range of cells types including adipocytes, osteoblasts, hepatocytes

Question 5

what are totipotent cells?

Answer 5

• are able to differentiate into any type of cell
  
  • in humans, this applies only to the zygote through the stage of the morula

Question 6

what are pluripotent cells?

Answer 6

• able to differentiate into any of teh germ layers (endo, ecto, or mesoderm), and can be obtained from the internal cell mass of the blastocyst

Question 7

what are multipotent cells?

Answer 7

• adult stem cells
• refers to the ability to differentiate into several types of cells within a relatively limited functional scope

Question 8

what are oligopotent cells?

Answer 8

• a stem cell that can only derive into a few types of cells

Question 9

what is apoptosis?

Answer 9

• programmed cell death

Question 10

what is the concept of operon?

Answer 10

• gene expression in prokaryotes
• they are relatively simple and mechanistic systems that allow a bacterium to respond to changes in its environment by increasing or decreasing the expression of certain genes as appropriate

Question 11

what is an operon under negative control?

Answer 11

• a repressor prevents transcription by binding to the operator (a sequence upstream of the first protein-coding region)

Question 12

what is an operon under positive control?

Answer 12

• an activator stimulates transcription

Question 13

both the lac operon and trp operon involve what kind of control?

Answer 13

• negative control, but differ in that the lac operon is inducible and the trp operon is repressible

Question 14

what is the mechanism of a negative inducible and negative repressible operon?

Answer 14

• inducible- the repressor is normally present and the genes are not expressed except under specific conditions
• repressible- the genes are usually transcribed, but transcription can be halted by the binding of the repressor in appropriate conditions

Question 15

• what is the mechanism of action of the lac operon?

Answer 15

• discovered in E. Coli but the basic idea is that E. Coli has the ability to emtabolize glucose, while the lac operon gives E. Coli the ability to metabolize lactose if it is present
• however, expressing the proteins necessary to metabolize lactose is energetically somewhat expensive, so it is advantageous for E. Coli to express that cellular machinery only when lactose is present, and more so when lactose is present but glucose isn’t which is what the lac operon does
• when no lactose is present, the repressor is bound to the operator and prevents RNA polymerase from transcribing the structural genes

Question 16

lac operon table summarized?

Answer 16

Lactose| Glucose| Repressor| Cap| outcome

No| yes| bound to operator| absent| no lactose metabolism

Yes| Yes| released from operator| Absent| weak lactose metabolism

yes| No| released from operator| Binds to CAP binding sequence| strong lactose metabolsim

Question 17

what is the mechanism of action of trp operons?

Answer 17

• contain genes for the synthesis of tryptophan, unfavourable for these genes to be expressed if tryptophan is present, so trp operon allows these genes to be expressed in the absence of tryptophan
• when tryptophan is absent, the repressor does not bind to the operator, and tryptophan synthesis proceeds
• when tryptophan is present, it binds to the repressor protein and causes it to bind to the operator, thereby inhibiting synthesis

Question 18

what are promoters in eukaryotes?

Answer 18

• many promoters are characterized by specific highly-conserved gene sequences
  
  • the TATA box is located about 30 bp upstream of the coding sequence, while the GC box and CAAT box are located roughly 10-150 bp upstream of the TATA box
  • they bind proteins that help recruit DNA polymerase to initiate transcription

Question 19

what are transcription factors in eukaryotes?

Answer 19

• proteins that regulate expression by binding to a specific DNA sequene through what is termed a DNA-binding domain
• once bound, they can recruit other regulatory proteins

Question 20

what are enhancers in eukaryotes?

Answer 20

• allow gene expression at even higher levels than usual in response to the appropriate stimuli
• do not have to be located in close proximity to the coding region of the gene like promoters
• enhancers bind transcription factors that twist DNA into a hairpin loop, brining distant regions into close proximity to each other

Question 21

what are silencers?

Answer 21

• regions of DNA to which transcription factors known as repressors bind and inhibit the transcription of a gene that they target

Question 22

what is the purpose of DNA methylation?

Answer 22

• a methyl group is added to cytosine or adnine
• generally thought to inactivate genes
• plays a major role in epigenetics

Question 23

what is the phenomenon of epigenetics?

Answer 23

• inheritable phenotypic changes involving mechanisms other than the alteration of the genome itself

Question 24

what is non-coding RNA?

Answer 24

• refers to RNA that is not translated into a protein ex. tRNA
• siRNA- short, double-stranded
• miRNA- single nucleotide strands incorporated into an RNA structure with a characteristic hairpin loop

Question 25

what is the function of siRNA and miRNA?

Answer 25

• silence genes by interruppting expression between transcription and translation

Question 26

what is the role of enhancer RNA (eRNA)?

Answer 26

• eRNA sequences are transcribed from enhancer regions of the DNA and appear to be associated with increased transcriptional activity

Question 27

what is a tumor?

Answer 27

• describes any abnormal proliferation of cells
  
  • benign tumors remain localized
  • malignant tumors can invade other organs and tissues in the body, in a process called metastasis

Question 28

what is tumor initiation?

Answer 28

• the first step in oncogenesus, involves changes that allow a single cell to proliferate abnormally
• tumor progression occurs as a cell develops the ability to proliferate even more aggressively

Question 29

what is the difference between oncogenes and tumor suppresor genes?

Answer 29

• oncogenes function to promote abnormal growth and poliferayion leading to cancer while tumor suppressor genes function to prevent tumorigenic properties

Question 30

what are proto-oncogenes?

Answer 30

• A gene involved in normal cell growth. Mutations (changes) in a proto-oncogene may cause it to become an oncogene, which can cause the growth of cancer cells.

Question 31

what is the process of recombination?

Answer 31

• Recombination is a process by which pieces of DNA are broken and recombined to produce new combinations of alleles. This recombination process creates genetic diversity at the level of genes that reflects differences in the DNA sequences of different organisms.

Question 32

what are vectors?

Answer 32

• DNA molecules used to carry genetic material into a cell where it can be replicated or expressed
  
  • 2 main types used are plasmids and bacteriophages

Question 33

what are plasmids?

Answer 33

• short circular DNA molecules that can replicate independently in bacteria

Question 34

process of recombinant DNA

Answer 34

1. Cut open the plasmid and “paste” in the gene. This process relies on restriction enzymes (which cut DNA) and DNA ligase (which joins DNA).
2. Insert the plasmid into bacteria. Use antibiotic selection to identify the bacteria that took up the plasmid.
3. Grow up lots of plasmid-carrying bacteria and use them as “factories” to make the protein. Harvest the protein from the bacteria and purify it.

Question 35

Bacteriphages as vectors are used for?

Answer 35

• larger sequences such as 15kb

Question 36

what is complementary DNA (cDNA)?

Answer 36

• a way to clone RNA sequences
• the first step is to synthesize a DNA copy of the RNA using reverse transcriptase (the DNA product is called cDNA because it is complementary to the template of RNA) and this can then be ligated to vector DNA

Question 37

what does the term transgenic refer to?

Answer 37

• refers to any organisms whose genome has been modified

Question 38

what are knockout organisms?

Answer 38

• those in which one or more genes have been disabled

Question 39

what is gel electrophoresis?

Answer 39

• a technique used to analyze nucleic acids by size
  
  • the principle of gel electrophoresis is to suspend charged macromolecules in agarose gel and apply an alectric field by applying a positive charge (anode) at one end and a negative charge (cathode) at another end (this is an electrolytic cell)
  • macromolecules will move in response to charge, but their movement is affected by size with larger molecules moving more slowly and smaller molecules moving more quickly
  • nucleic acids with a high degree of negative charge due ti the presence of many phosphate groups will migrate towards the positively charged anose
• following electrophoresis, the DNA is stained with fluorescent dye and photographed
• molecular weight size markers (ladders) are fragments with known sizes that can be run parallel to the DNA of interest in an electrophoresis experiment to identify the approximate size of the DNA fragments being analyzed

Question 40

what is hybridization?

Answer 40

• the ability of single-stranded DNA (or RNA) to form base pairs with a complementary sequence
• important step in polymerase chain reaction (PCR)
• can also be used to identify target sequences by using a hybridization probe which is a specific DNA or RNA fragment that is known to the researchers that can be labeled radioactively. if it is added to a sample of DNA or RNA being studied and the sample is heated to generate single-strand DNA or RNA, then the single-stranded radiolabeled probes will anneal with complementary sequences (helps determine the presence of the sequence of interest in the DNA that is tested)

Question 41

the principles of electrophoresis and hybridization are combined in?

Answer 41

• blotting techniques
  
  • southern blotting is used to identify DNA sequences
  • Western blotting is used to determine protein sequences
  • Northern blots are used to identify RNA sequences

Question 42

how do blotting techniques work?

Answer 42

• The molecules of interest (DNA, RNA, or proteins) undergo gel electrophoresis to separate them by size, and then are transferred to a nitrocellulose membrane that can be heated, at which point probe analysis can be performed (the process is slightly different for western blots, where antibodies are used instead of DNA/RNA probes)

Question 43

what is DNA microarray?

Answer 43

• A DNA microarray (also commonly known as DNA chip or biochip) is a collection of microscopic DNA spots attached to a solid surface. Scientists use DNA microarrays to measure the expression levels of large numbers of genes simultaneously or to genotype multiple regions of a genome.

Question 44

What is the Sanger method or the dideoxy chain termination method?

Answer 44

• uses dideoxynucleotides to terminate synthesis and electrophoresis to analyze fragment size

Question 45

what is the process of PCR?

Answer 45

• Polymerase chain reaction (PCR) is a common laboratory technique used to make many copies (millions or billions!) of a particular region of DNA.

The basic steps are:

1. Denaturation (96°C96°C96, °, start text, C, end text): Heat the reaction strongly to separate, or denature, the DNA strands. This provides single-stranded template for the next step.
2. Annealing (555555 - 656565°C°C°, start text, C, end text): Cool the reaction so the primers can bind to their complementary sequences on the single-stranded template DNA.
3. Extension (72°C72°C72, °, start text, C, end text): Raise the reaction temperatures so Taq polymerase extends the primers, synthesizing new strands of DNA.
   * This cycle repeats 252525 - 353535 times in a typical PCR reaction, which generally takes 222 - 444 hours, depending on the length of the DNA region being copied. If the reaction is efficient (works well), the target region can go from just one or a few copies to billions.
   * Uses Taq polymerase (heat stable enzymes)

Question 46

what is the role of restriction enzymes?

Answer 46

• cut DNA ar specific points, leading to blunt and sticky ends that can be recombined