Thank you, and good luck next year!

Thank you, T04-T07 students, for all the hard work you put in, and for all your interesting, funny, creative comments and questions!

Have a great Summer break!


All the best,

Pam

PS: I am still trying to find an aswer for that sperm cytoplasm P element question...
and for those who were interested in finding out what type of smart they are, have a look at
http://www.berghuis.co.nz/abiator/lsi/lsiframe.html.

A couple of things

1) Nested enzymes
There is no such thing…but there are such things as nested restriction site recognition sites and molecular biologists who make up confusing expressions.
The idea is that,sometimes, when you cut a vector with enzyme I and the DNA to be inserted with enzyme II, and you join them together, you may re-generate the recognition site for enzyme I or enzyme II (depending on the situation) or generate a site for another enzyme altogether (enzyme III). In some cases, the outcome of the rejoining procedure depends on what nucleotide is present in the insert just adjacent to the restriciton site for enzyme II.
What may be confusing is that you may be cutting the vector with enzyme I, cutting the genomic DNA with enzyme II, and after rejoining you obtain 3 clones of interest (containing different genomic fragments), and clone I may have regenerated one site for enzyme I at one junction, while clone II may not have regenerated any site, and clone 3 may have regenerated the site for enzyme I at both junctions.
Just treat each clone as a separate entity!


2) Drugs and cell signalling : here’s a fun site if you need to take a short break from studying: go to
http://learn.genetics.utah.edu/units/addiction/drugs/mouse.cfm

3) Please remember : you won’t need to memorize any gene names or functions. Just keep in mond some examples of mechanisms, e.g. gradients of proteins/RNAs, cell-to-cell signalling, transcriptional regulation, post-transcriptional regulation (e.g. inhibition of translation), etc.

4) tomorrow is review day. Try to come in with a list of specific questions that you have....

development stuff

On P elements and chapter 17

More questions

On RFLPs, chromosome walking, etc!

More questions

CLICK ON THE QUESTION TO ENLARGE

markers question

extra help session

Next Friday, the 13th of July, I'll be in Buchanan, room D 322 from 1.45 to 4.30 pm. Instead of a tutorial, we'll have a Q&A/help desk session. the idea is that you come in with your questions, or with some unsolved problems and I'll help you out with them.

Tomorrow, Wednesday, I'll be in D322 for at least 30 min after the end of the tutorial.

Practice questions on RFLPs, VNTRs, and molecular markers in general will be posted here in the next 24 hours!

Cheers,

Pam

post-midterm thoughts

I am very sorry about how people are feeling. I feel terrible as a TA, for various reasons, and I can only emphasize with you. On the other hand, I commend you all for sharing your thoughts and feelings-keep them coming! It's very informative, and it's good for people to see that they "are not alone".

Now, here is the tricky part: do you really want to do something tangible about it? I can point out what I don't like about the exam, but this is very unlikely to have any effect, if past experience is a reference. Each one of you can go complain bitterly about how "unfair" it was, which also is not very likely to have any consequences.
Or, we can look at is as scientists, make a coordinate effort, base all arguments on facts, and come up with an objective conclusion and possibly a 'suggestion for improvement' or 'possible solution'. That is much more likely to have an effect.

I don't think anyone will be able to look at this situation objectively enough for the next day or so, as it was a very upsetting experience. When you feel a bit better, you can think about, and maybe quickly write down the reasons why you think that question 1, or 2, or 3 were "bad", and what you would do to make them better. You can also think about how much information, and how much practice did you have on the lac operon? How much about homologous recombination in bacteria? How much about ligations and restriction mapping?
I'll be at Monday's tutorial 15-20 min in advance and I'll stay later, so we'll have time to discuss the matter and decide what to do about it.

Cheers,

Pam

PS: for the record, I think that question 2 was an old question from Dr Beatty (the formatting looks exactly like his, and not at all like Craig's)

A few answers

I see that there is some action going on here, nice job everyone!

Let's try to address your concerns in some sort of organized way:
-I don't think Omid's mock exam is available yet, I tried to download it too and it wasn't accessible.
- your midterm predicitons seem pretty sensible to me: lac operon-type (prokaryotic gene regulation) stuff, restriction mapping and a possible cloning project....sounds good! But please realize that I don't know what Craig is planning on doing.

Cloning project: we've seen, so far, 3 basic approaches (functional complementation-based cloning, cloning based on sequence homology, and you guys did cloning based on differential expression/expression profiles in class). Note that not all these approaches need a probe! The first one, for example, is based on restoring a function, and you don't need any probes to check for a function.

About the outlined cloning procedure:
-first of all, you need to know what type of gene you are trying to clone, and what information is available!
- you need to say where your DNA, that you partially digest, comes from (a bacterium? plant? fungus? Is it genomic DNA? Is it subcloned DNA? etc)
-you ALWAYS want a selectable marker in your plasmid, typically a drug resistance. People do not use the LacZ+ as a selectable marker because it complicates the procedure (you can't use standard media, etc). The lacZ gene on the plasmid is used for a different thing...we can go over this later.
-If you are using colony lift hyb. to find the colony that contains our gene of interest, that means you already know the gene sequence....so why wouldn't you skip the whole procedure and just PCR it up from genomic DNA? You need to be very clear about the goal of your project, what you have at yoour disposal and what you don't. Different situations require different procedures!
- you don't run Southern blots on bacterial colonies...you would need 100s of gel lanes, which means dozens of gels!!

NOTE that, in all cases, once you have identified a clone that contains your entire gene, you don't need to do anything to it; you got it. To amplify it you just grow up your bacterial culture that contains the plasmid with your entire gene of interest, the next day you do a plasmid prep from that culture, and there you have tonnes of your gene of interest, conveniently inserted into a plasmid. If you want to cut it out it's really easy because you can use restriction enzymes!

See you in class!

Pam

QUESTIONS THAT CAME UP!

Thank you very much for taking the time to ask all these questions in an organized way. I tried to do my best to clarify...let me know if it helps (or if it doesn't!)

Cheer,

Pam