Thursday, July 12, 2018

CRISPR Cas9 for Dummies

From Synthego
I have to admit that the ongoing revolution in gene editing can be more than a bit confusing -first there were transgenics, then targeted mutations, conditional mutations, zinc fingers and TALENs ...   Most recently we have CRISPR Cas9.   Because it looks like this method really is the holy grail of gene editing, it's time for a few basics (even for those of us clinicians who qualified when gene editing wasn't even a gleam in some researcher's eye).



1. In what categories of organism was the CRISPR Cas9 system discovered?
2. What was its "native" purpose?
3. What does CRISPR Cas9 stand for?
4. If we say that transgenic mice are created by inserting genetic material into the male pronucleus of the oocyte by microinjection,  and gene-targeted (knockout) mice are created by inserting genetically modified ES cells into the blastocyst by microinjection, how are CRISPR Cas9 mice created?
5. True/False: CRISPR Cas9 has been used to make genetic modifications in plants, multiple animal species and human genetic material.
6. From which organism is the most commonly-used type of Cas9?
7. What 2 functional components constitute the CRISPR Cas9 system?
8. True of false. The CRISPR Cas9 system relies upon the host cell-repair system to function.
9. True or false: CRISPR Cas9 can be used to knock out genes, to knock in genes and to study the genome wide function of genes and non-genomic DNA through interference.
10. True or false: to date CRISPR Cas9 improves upon existing methods for gene modification in all areas.


ANSWERS POSTED HERE






Monday, June 25, 2018

Q32: Ferrets and their kin

Image from Fitzroy hospital
1. Unbelievably, ferrets have been domesticated for more than 2000 years.  They belong to the subfamily _________ which contains 5 subgenera - name the subgenera and their common names.

2. The ferret is being used to replace the cat in which (general) types of experiment?

3. What special light considerations are needed to maintain successful breeding in ferrets in the lab?

4.  What features of the gastrointestinal tract are reflective of the fact that they are strict carnivores?

5. What features of their reproductive cycle make them good research subjects for study of the HPA axis?

6. Which organism commonly causes acute abdominal distension with dyspnoea and cyanosis in weanlings subjected to sudden dietary changes or overeating?

7. Which organism is widespread in ferrets, often asymptomatic, but can cause gastritis and peptic ulcers?  How would you treat it?

8. Which organism is commonly associated with proliferative bowel disease?

9.  What is the most serious viral disease of ferrets causing almost 100% mortality?  Describe the phases of this disease.

10.  What viral disease would you suspect if the principal signs were hypergammaglobulinemia and chronic weight loss?  What is the most consistent histological finding with this disease? Is there a vaccine available?

11. Which disease is a zoonosis and has been used extensively to model the disease in humans?

12. What clinical chemistry abnormalities are found in 70% or more of ferrets with pregnancy toxemia?  What percentage of ferrets with this condition will be anemic?

13. What is the treatment for hyperestrogenism?

14.  Which 4 types of cancer account for most ferret neoplasms?

15. In aging ferrets, what common condition presents with weight loss, lethargy and anorexia?

ANSWERS POSTED HERE


Friday, June 8, 2018

Q 31. Other rodents

This week we will cover some of the less well known species that have some interesting and unique features that are useful for research.

1.  Match the following rodents with their suborders:


Suborders:

  1. Sciuromorpha
  2. Hystricomorpha
  3. Myomorpha
  4. Castorimorpha

Rodents
Chinchilla, kangaroo rat, prairie dog, hamster, naked mole rat, mice, guinea pigs, pocket gophers, voles, degus, squirrels, gerbils, grasshopper mouse

2. Why is there a ban on import of exotic African rodents for the pet trade?  Which of the rodents was involved in the incident that resulted in this ban?

3.  What is the genus and species of the kangaroo rats?  List 3 uses in research and indicate what features of the kangaroo rat make them useful for these models.

4. Which of the rodents listed in Q1  hibernate?

5. What are the genus and species of the 2 deer mice?   List 2 major N American zoonotic diseases for which they are a vector.  Bonus question: how many other zoonotic diseases can you name that they can be a vector for?

6. Which rodent (genus species, common name) is the natural host for Venezuelan hemorrhagic fever?

7. How do you collect blood from the cotton rat? (hint:it's not the usual site)

8. Two of these rodents are used to study dental disease.  Which rodents and which diseases are they used for?

9. Which rodent is the model for Zollinger Ellison syndrome?  What is Zollinger Ellison syndrome?

10. Which rodent is carnivorous?

11. Identify these rodents (common name, genus and species) - as a lot of them look similar- I've provided some hints!

Indifferent to photoperiods...
Um... white..?

Resistant to venom and eats scorpions


Look at the ears! ( and can't make up it's mind about shiny objects)
hint - it's not a mouse
check out that coat!

This one should be obvious
Look at those feet!
Federally endangered!
name means small ears


How did you do?

ANSWERS POSTED HERE



Monday, May 7, 2018

Q 30: Hamster happenings




Today's questions are about a rodent we don't see so many of in our animal facilities, but they surely have some interesting characteristics...  and here are some questions to perhaps make you check out the blue book!

1. There are 4 genera of hamsters we use in research:
Name the genera – and might as well name the species too…
  • Which is the smallest? 
  • Which one does NOT hibernate and which one is a TRUE hibernator?


















2. We know that hamsters have ‘special’ cheek pouches:
  • What is special about them and why are they that way?
  • What are they often used for?




Golden (Syrian) Hamsters

3. True or false?
  •  They hibernate
  • They need to fatten up before hibernation
  • If they wake during hibernation they won’t go back into hibernation

4. What is different about the anatomy of the stomach in hamsters compared to other rodents?

5. What characteristics of their pulmonary system makes them a good model for smoke inhalation studies?

6. How are their oocytes used in a biomedical/clinical setting?

7. What is the cause of “wet tail” in hamsters?

8. Tyzzers disease in hamsters has a triad of pathologic signs: what are they?

9. What is unique about the hamster Helicobacter aurati (compared to mouse and rat Helicobacter species)

10. At what ABSL must hamsters be kept if they are infected with lymphocytic choriomeningitis? Is this the same as in mice?

11. Hamster parvovirus is nearly identical to which mouse parvovirus?

12. Name three differences (2 anatomic, one epidemiologic) between the 2 cestodes that hamsters are susceptible to.. (Rodentolepis nana and Hymenolepis diminuta)

Other Hamsters


13. Which species is the most radioresistant and develops spontaneous diabetes mellitus?

14. Which species is highly susceptible to oncogenic viruses? – can you name (3 or 4) oncogenic viruses?

15. In general, when breeding hamsters, the male should be taken away for raising the young. Which hamster species is the exception and you should leave the male to help raise the young?

Answers posted here



Monday, April 2, 2018

Zebrafish 101



Zebrafish are an increasingly common sight in or near our research facilities. How much do you know about keeping fish (and zebrafish in particular)?



1. Which characteristics of zebrafish development make them so attractive for studying embryonic development?

2. What is the genus and species and native location

3. Zebrafish water quality parameters should be:
a. pH
b. temperature
c. conductivity
d. hardness

4. True or false, ammonia , nitrite and nitrate are very toxic to fish

5. Why should the temperature of the tank room be maintained slightly higher than tank water?

6. When retrofitting a fish facility into a room on an upper floor – what construction detail should not be overlooked?

7. What is the best method to control excess algae in the aquarium

8. What is the commonest cause of fish becoming ill due to infectious disease?

9. Which Mycobacterium species are commonest in aquarium fishes? Can they infect people?

10. What are the commonest bacterial infections?

11. Which is the commonest nematode and what is the characteristic feature of its eggs?

12. Which is the commonest flagellate skin disease and what characteristic behavior is seen with infected fish?

13. What is the commonest fungal disease caused by and is it usually primary or secondary? What is the prognosis?

14. What is the commonest ciliated protozoan parasite of skin?

15. Which ciliate protozoan parasites infect gills? Are they common in the lab?

16. When treating a tank with antibiotics, what should be done with respect to external filtration? Which drug could be photoinactivated?

17. Which diseases can be treated by salt water immersion and raising tank temperature?

18. Which disease requires depopulation and intensive environmental treatment with bleach to prevent reintroduction from the environment?

19. Malachite green is a popular treatment for fish infections, which should not be used in the laboratory. Why not?

20. Not to belabor the point, but what is the commonest cause for sick fish?

ANSWERS POSTED HERE




Monday, March 5, 2018

Q 28. What do you know about humanized mice?

from the institute of  molecular and cell biology
These super-expensive mice are increasingly arriving in our facilities as the latest tool in combating human cancers and infectious and autoimmune diseases.  Here are some questions relating to key facts and concepts you might need to know.

1.  What was the key additional mutation that allowed immunodeficient mice to be engrafted with components of the human immune system and tissues?


2. What are the 3 immunodeficient mouse strains that together with this mutation are used as models to engraft human tissues. (gene names and symbols please!)

3.  Which model (common name) is engrafted by injection of peripheral blood leukocytes?  What are 2  disadvantages of this model?

4. Which model (common name) is engrafted by IV or intrafemoral injection of human SCID-repopulating cells?  What is the advantage of this model over the previous one?

5. Which model (common name) uses human fetal tissues and what is the deficiency of this model?

6. Humanized mice are frequently used to study HIV.  Which mouse model is most commonly used and why?

7. Epstein Barr virus is studied using humanized mice. EBV affects 90% of humans worldwide. Which cells are transformed by EBV and which immune cell is normally responsible for eliminating these transformed cells? List immunological and autoimmune disorders that can result from EBV if these cells are not removed.

8. Most humanized mouse models have used human cell lines for engraftment, however unlike in patients, cell lines and the environment they grow in is homogenous. In the next step forward, patient cancer cells are being grown in humanized mice. Which of model (of the 3 types above) is required for growth of human patient cancer cells? What is the common name of this model?

9. List 3 (or more) remaining problems to be solved in development of future humanized mouse models.

ANSWERS POSTED HERE

Monday, February 5, 2018

Q27: Spontaneous Diseases in Common Mouse Strains

image from clipartbarn
Why should you care?  Well, it's not unheard of for our investigators to suspect that pathology or behavior they discover is related to a mutation,  when in fact it's a feature of the background strain. 

So here is an eclectic collection of questions relating to our common inbred strains...

1. Which strain has long been used as a source of ES cells for targeted mutations and why is this a problem?

2. List 2 strains that have mutations in the dysferlin gene and what condition (s) results?

3. Which strain is known for has a high incidence of thymic T cell lymphoma and what is the genetic reason?  Which immunodeficient strain is also has a high incidence of T cell lymphoma?  Which strain has a high incidence of B cell lymphomas?

4. Which strain has a high incidence of cardiac abnormalities including epicardial mineralization and cardiomyopathy and is widely used for infectious disease studies?

5. Which 2 strains with high and low mammary tumor incidence (respectively) were developed from an A albino female to a DBA male?

6. Which strain historically had a very high incidence of mammary adenocarcinomas due to MMTV?

7. Which strain is known for its resistance to many infectious agents but susceptibility to mouse adenovirus 1 and encephalomyelitis?

8. Which induced disease  model is the DBA mouse known for and what is the human condition?

9. List 2 strains susceptible to seizures

10. List 2 strains that exhibit acallosity


ANSWERS ARE POSTED HERE