Microbiology New Material

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1

Describe lac operon and repressor gene.

Lac operon is a series of 4 genes.

The repressor gene blocks the other 3 genes access if repressor is there. The RNA polymerase can't get through and copy.

If lactose comes into enviroment of E.coli, lactose will be in charge of taking the repressor off. It's the sugar that pulls the door open.
1. permease transports lactose in
2. B-galactosidase

In order to carry out glycolysis, we need glucose from the splitting of lactose from B-galactosidase which yields glucose and galactose.

2

What are the 2 types of operons?

1. Inducible operons (ex. lac operon) usually control catabolic pathways. They break down lactose to get energy (energy generation). The thing that dictates whether it's on or off is the presence of substrate of pathway.

2. Repressible operons: anabolic pathways; energy consuming; presence of product of pathway. Ex. trying to build hair (costs body energy) consuming energy to build hair. If you have hair, then it turns off. Vise versa.

3

Explain the regulatory decisions of lactose.

Is lactose present?

If yes, Is preferred carbon and energy source present? (glucose)
If yes, don't synthesize enzymes (gene/operon off)
If no, synthesize enzymes (gene/operon on)

Is lactose present?

If no, don't synthesize enzymes (gene/operon off)`

4

What explains the diauxic growth of E.coli and describe it?

The lac operon explains the growth of E.coli w/ a mixture of lactose and glucose. A short lag in growth is present while the bacteria synthesizes the enzymes needed for lactose use.

1st growth: using glucose
stationary phase: time it takes for E.coli to realize glucose is gone and turn on gene to use lactose
2nd growth: breaks down lactose and uses glucose again

5

What bacteria produces insulin? Describe how it does this.

E.coli. It makes it b/c we gave it a human gene. We all need it but diabetes folks have defective genes so they have to get it injected. Pig insulin and cow insulin work in humans. This was a problem b/c people could build a resistance to it and it wouldn't work.

6

What are 3 ways of getting growth hormone?

1. cow they made can make human growth hormone
2. It's now made in genetically modified bacteria
3. New way of proposal: give cow gene, it expresses gene, and releases it in her milk.

7

What are the desirable features in microbial cloning hosts? Why do we use bacteria to use human insulin?

Rapid overturn, fast growth rate (can't do this to human cells) (cancer cells even slower than bacteria)
Can be grown in large quantities
Nonpathogenic
Genome (genes) that is well mapped
Capable of accepting plasmid
Maintains foreign genes through multiple generations
Secrete increased yield of proteins from expressed foreign genes

*E.coli does all of the above EXCEPT it doesn't secrete proteins

8

What are the only prokaryotes?

bacteria

9

What is the most commonly used microbe?

E.coli

10

What are two tools needed for cloning?

1. restriction enzyme- made by bacteria; cuts DNA
2. DNA ligase-seals

We first cut DNA to get gene and cut plasmid
Insulin gene slips into plasmid where it was cut.. DNA ligase seals human gene and plasmid
Recombined plasmid
Cloning of the insulin gene

11

What are 3 reasons why we clone?

1. protein product
2. study a gene
3. bacteria itself is what you want

12

Describe the restriction enzyme.

Come from bacteria; cuts DNA. Diff. bacteria produce diff. restriction enzymes. Specific for certain sequences in DNA. You choose which enzyme based on what sequence you won't to cut.

13

Why do they make enzymes to cut DNA?

bacterial immune systems- when they get infected by viruses they inject DNA material in you. So, if DNA comes in from virus we have a way to get rid of that sequence of DNA w/ enzyme. It is a protection from foreign DNA.

14

Know the ECORI recognition site.

GxA-A-T-T-C
C-T-T-A-AxG

Turns into complementary tails. You want tails of plasmid gene to equal tails of insulin gene.

DNA ligase seals the gaps

15

What is a commonly used plasmid?

pUC19- E.coli plasmid that is small. High copy # (copies itself) Temperature dependent copy. 42C makes >200 copies. About 75 per cell @37C.

16

What corrects human growth defiencies in children?

HGH

17

Describe the Epidermal growth factor (EGF)

Heals wounds, burns, ulcers, produced by E.coli. Important roles in normal development. Today, we can make this.

18

What is desmoteplase?

An anticoagluent gene in vampire bats. They put this gene in E.coli so it produces it.

19

What bacteria produces pig and cow growth hormones?

E. coli

20

How do we get genes in plants?

We take DNA and coat it w/ something heavy enough to shoot it out of a gun. They shoot particles into a seed b/c plants have thick cell walls. We stabilize genes by getting them into chromosomes. The gene will CROSS-OVER into chromosome.

21

What is transgenesis?

genes from an unrelated species incorporated into a plant or animal's germ line.

22

what bacteria is toxic to caterpillars?

Bacillus thuringiensis. Monsanto decided to put the bacteria's toxic gene into plants. Another example they did to show people gene transplanting works is they gave a tobacco plant a lightening bug gene so it lit up. They also have made blue roses.

23

How do they get genes into animals?

Use a needle to inject gene into an egg. they are now using a virus injected w/ gene to get into the egg. Viruses know how to get genes and insert them into chromosomes properly. The virus is disabled so it can't reproduce and can't infect.

24

What is GFP?

(Green fluorescence protein). It is a color reportor gene. You cut this gene and stick it w/ the gene you want to be expressed. If you see green color then you know the gene is there too.

We have put it in mice and if the baby mice are green, then we know the gene they want is expressed.

Zebra fish are the most used since they are transparent to view internal organs.

25

What animla did they use the GFP for to study Huntington's disease?

used monkey's

26

What are distinguishing characteristics of viruses?

1. obligate intracellular parasite
2. incapable of independent metabolism
3. they are filterable (much smaller than bacteria)
4. they possess only a SINGLE type of nucleic acid, either DNA or RNA.

27

T or F. Virues shapes are more diverse than bacteria. They are very distinctive looking.

T.

28

What are bacterialphages?

virus infecting bacteria. Virus can even infect plants. They can be good thought. Deli's can spray meat w/ viruses and the they will inject any bacteria that is in the meat.

29

How do we know we have a virus? What are the 3 commom ways to see them? What's 2 more not so common ways.

Humans can't grow viruses on streak plates or see them under a microscope.

1. some viruses are detected by signs and symptoms of patients.
2. take human cells and see if they look normal or abnormal (abnormal may be caused by virus)
3. check for antibodies. If it's an elevated count, may have a virus.

4. Stick sample under electron microscope (hospitals don't really do this)
5. DNA analysis (expensive; not developed well)

30

T or F. Diff. viruses do diff. things to cells.

T. Herpes virus causes cells to FUSE TOGETHER and ROUND UP.

31

What can viruses cause?

1. lytic infection (virus multiplies and kills host cell immediately) EX. smallpox, influenza

2. persistent infection (virus lives in cell long period of time) EX HIV

3. Latent infection (virus resides in cell but produces no viral particles) EX chicken pox

4. Cancer-causing infection (virus transforms cell into a cancer cell that becomes a seed for tumor) EX HPV (causes warts)

32

Describe HPV

Animals can get it
Incubation period: 3 months
Symptoms: may have no symptoms; warts of genitals
Asymptomatic person can transmit disease

33

What types of HPV cause 90% of genital warts?

Types 6 and 11

34

what types of HPV cause 70% of cervial cancer?

Types 16 and 18

35

Know that the Nobel peace prize was won by a guy who found the virus that causes cervical cancer

...

36

What percent of 20-24 yr olds were HPV-positive?

45%

37

What is gynocologist Dr. Lori Stafford's quote about HPV

"It's (HPV) the common cold of gynocology"

38

T or F. HPV may enter through genital scrapes. Virus may inject its DNA into your chromosomes, if so you will never get rid of it and may develop cervical cancer.

T.

39

Do we have a vaccine against HPV?

Yes, Gardasil