AP Bio Unit 6B Shigella Lab+ Retroviruses+ CED + CRISPR

Phenotype determined by genotype CED: Genetic information in retroviruses is a special case and has alternate flow of information” from RNA to DNA, made possible by

reverse transcriptase, an enzyme that copies the viral RNA genome into DNA. This DNA integrates into the host genome and becomes transcribed and translated for the assembly of new viral progeny.

Mutations contribute to variation subject to natural selection CED: Changes in genotype may affect phenotypes that are subject to natural selection. Genetic changes that

enhance survival and reproduction can be selected for by environmental conditions

Natural Selection Genetics CED: The horizontal acquisitions of genetic information primarily in

prokaryotes via transformation (uptake of naked DNA), conjugation (cell to cell transfer of DNA), and transposition (movement of DNA segments within and between DNA molecules) increase variation.

Natural Selection Genetics CED: Related viruses can combine/recombine

genetic information if they infect the same host cell

Natural Selection Genetics CED: Reproduction processes that increase genetic variation are

evolutionarily conserved and are shared by various organisms

Genetic engineering techniques used to analyze and manipulate DNA/RNA CED: Electrophoresis separates molecules

according to size and charge

Genetic engineering techniques used to analyze and manipulate DNA/RNA CED: During Polymerase Chain Reaction (PCR),

DNA fragments are amplified

Genetic engineering techniques used to analyze and manipulate DNA/RNA CED: Bacterial transformation introduces

DNA into bacterial cells

Genetic engineering techniques used to analyze and manipulate DNA/RNA CED: DNA sequencing determines the

order of nucleotides in a DNA molecule

Characteristics of VIRUSES: non cellular, non living structures

contain a protein coat called a CAPSID

nucleic acid containing DNA or RNA (DS or SS)

capable of “reproducing” only when within a HOST cell

Key variables to classify viruses

 DNA or RNA? 2. Is the genetic material single stranded (SS) or double stranded (DS)? 3. What is its shape/structure? 4. What is its reproductive strategy? Lytic? Lysogenic?

Lytic - Steps: “short and sweet”

1. Viral information is injected into the host cell.

2. The protein machinery is hacked.

3. The machinery churns out viral proteins/parts for assembly.

4. Lots of viruses replicate inside the cell.

5. The cell lyses and the viral material is released.

Lysogenic - Steps: “sneaky, idle, more dangerous”

1. Viral information is injected into the host cell.

2. The viral information is turned into a form that allows it to be integrated within the host’s DNA.

3. The host cell replicates like normal.

4. One day it is triggered, LARGE VIRAL RESPONSE OCCURS- the lytic cycle begins (view steps above).

ex. HIV

HIV most common example of a

retrovirus- and lysogenic cycle

(retrovirus) RNA—→ DNA—→RNA—→Protein

When a retrovirus infects a host cell,

it injects its RNA and an enzyme called reverse transcriptase into the cytoplasm

Retrovirus- - A type of virus that contains the enzyme reverse transcriptase, an enzyme that can turn a piece of

single-stranded RNA into complementary DNA. That DNA is then integrated into the host’s DNA. This lysogenic strategy makes the virus much more stealthy and dangerous.

CRISPR- gene editing tool

It is found naturally in bacteria and works as an immune system. CRISPR can be used in ALL organisms.

Steps: 1. Virus invades a bacterial cell.

2. Cas proteins cut out pieces of viral DNA and it is integrated into the bacteria’s CRISPR region.

3. Viral DNA is transcribed into RNA where it binds to Cas9.

4. Cas9 latches onto any free-floating genetic material and destroys any viral material.

CRISPR in humans - Steps:

1. A guide RNA is made that matches the gene that has to be edited out.

2. With this information, Cas9 proteins cut out the unwanted genes.

3. Nonhomologous end joining and homology directed repair helps fix the DNA. The unwanted gene is gone and is replaced with something else.

Why is CRISPR used in humans? -

Are there ethical issues regarding the use of CRISPR? - Yes.

To edit out unwanted genes (like the ones that cause genetic disease).

Nonhomologous end joining:

Nucleases fix DNA by trimming off the broken DNA ends and gluing them back together.

Homology directed repair:

A second DNA template is made to guide DNA repair for CRISPR.

Why is it important to have clear information about everything the affected people ate?

It helps narrow down the list and brings us closer to the source of the infection.

What was the food item responsible? -

The 5-layer bean dip - This is because 5/6 affected people ate the bean dip. The other person didn’t eat the bean dip and either got sick from eating too much or from possible cross-contamination.

How do you know? -

The cheese layer had banding patterns that matched the SRS and +LC banding patterns.

MIM - Marker -

Tells us what each band means.

-C - Negative Control -

Contains no banding. If there is banding, that means there is contamination. 

+LC - Positive Control -

Contains banding (the different layers will have the same banding patterns as +LC). +LC tells us if the gel worked or not.

SRS - Shigella Reference Standard - Used for

identifying Shigella (our contaminated layer will have banding patterns that match both this and the +LC).