A (virtual) place for BIOL335 (2008) people to ask questions, answer questions, make requests, comment, wonder, discuss, complain and rejoice.
Thursday, January 31, 2008
Questions on lac operon and bacterial gene regulation
Please post here all your questions and requests regarding lac operon, problem set 2, chapter 10, and other topics relating to regulation of gene expression.
10 comments:
Anonymous
said...
Hi Pam,
If you had this: crp-I-P+O+Z+/crp-I+P-O+Z+ in the presence of both glucose and lactose. Would you get low B-gal activity(+), high B-gal activity(++) or no Bgal activity(-)?
If you had I-O+Z+Y+/I+O+Z-Y-, in the presence of the inducer would bgal and permease be (induced or constitutively made?) what is the difference between the two terms? Doesn't constitutively mean "always made" so say you had an Oc mutation and a P+ and a Z+ wouldn't this mean that b-gal would be constitutively made?
If you had: crp-I-P+O+Z+/crp-I+P-O+Z+ in the presence of lactose and glucose you'd get what I call "100" B-gal activity (in the absence of glucose you'd get 1000, in the absence of lactose you'd get 0). See, if you are crp-, then having glucose or not does not make a difference: you can never have "1000" B-gal.
If you had I-O+Z+Y+/I+O+Z-Y-, in the presence of the inducer you would make both B-gal and permease. In the absence of inducer you won't make them. Hence, in the presence of the inducer they will be induced.
Different instructors may use "constitutive" in different ways. I believe that you are absolutely correct, and "constitutive" means "always made" (hence the superscript "c" for the lacO[c] mutation!). However, you may come across people or books or handounts who use "constitutive" to mean "basal level", or "not fully induced". Stick with your definition-it's the correct one!!
I hope that I won't confuse you rather than clarify things....I'll try my best.
If it is P-, there's no expression (assuming that the mutation is a null mutation-if you have never heard of null mutations ignore this and take it as a yes, P-=no expression at all).
If everything works and there's no inducer, you may get a super low level of expression (like 1 unit of B-Gal).
If there's no cAMP, but there is inducer, you get a pretty good expression, like about 100 units of B-Gal. If there's no cAMP and no inducer, then you get 1 unit.
As a comparison, if you have lots of cAMP and inducer, you'd get 1000 units of B-Gal.
(If your questions come from trying a practice probelm that dealt with crp- mutants and talked about the need for cAMP in order to have maximum expression, don't worry-that question refers very specfically to materail that was taught that term).
Yes, specifically engineered bacteria will turn blue even without inducer (the typical E.coli lab strains have their lac operon deleted, and partially complemented in trans by bits and pieces in plasmids, and they may or may not have the gene encoding the lac repressor, so they'll be able to turn blue).
Don't worry about it, really. A few molecules of B-Gal will be able to make some blue colour, in some cells, and the blue colour is very visible even if only a few cells in a colony make it. But I promise you that that kind of blue colour is very different frokm the nice, strong blue that you get with induction!
Yes, specifically engineered bacteria will turn blue even without inducer (the typical E.coli lab strains have their lac operon deleted, and partially complemented in trans by bits and pieces in plasmids, and they may or may not have the gene encoding the lac repressor, so they'll be able to turn blue).
Don't worry about it, really. A few molecules of B-Gal will be able to make some blue colour, in some cells, and the blue colour is very visible even if only a few cells in a colony make it. But I promise you that that kind of blue colour is very different frokm the nice, strong blue that you get with induction!
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10 comments:
Hi Pam,
If you had this: crp-I-P+O+Z+/crp-I+P-O+Z+
in the presence of both glucose and lactose. Would you get low B-gal activity(+), high B-gal activity(++) or no Bgal activity(-)?
Thanx!!!!
Hi Pam,
If you had I-O+Z+Y+/I+O+Z-Y-, in the presence of the inducer would bgal and permease be (induced or constitutively made?) what is the difference between the two terms? Doesn't constitutively mean "always made" so say you had an Oc mutation and a P+ and a Z+ wouldn't this mean that b-gal would be constitutively made?
thx.
If you had:
crp-I-P+O+Z+/crp-I+P-O+Z+
in the presence of lactose and glucose you'd get what I call "100" B-gal activity (in the absence of glucose you'd get 1000, in the absence of lactose you'd get 0).
See, if you are crp-, then having glucose or not does not make a difference: you can never have "1000" B-gal.
Cheers!
Pam
If you had I-O+Z+Y+/I+O+Z-Y-, in the presence of the inducer you would make both B-gal and permease. In the absence of inducer you won't make them. Hence, in the presence of the inducer they will be induced.
Different instructors may use "constitutive" in different ways. I believe that you are absolutely correct, and "constitutive" means "always made" (hence the superscript "c" for the lacO[c] mutation!). However, you may come across people or books or handounts who use "constitutive" to mean "basal level", or "not fully induced".
Stick with your definition-it's the correct one!!
I am sorry about the confusion!
Pam
i have something to clear up
is the only time the lac operon not expressed totally (not even low) is when it is P-
even when the repressor works and there is no inducer, there is still low expression
and even when there's no cAMP to bind to the positive regulator, there is still low expression?
I hope that I won't confuse you rather than clarify things....I'll try my best.
If it is P-, there's no expression (assuming that the mutation is a null mutation-if you have never heard of null mutations ignore this and take it as a yes, P-=no expression at all).
If everything works and there's no inducer, you may get a super low level of expression (like 1 unit of B-Gal).
If there's no cAMP, but there is inducer, you get a pretty good expression, like about 100 units of B-Gal.
If there's no cAMP and no inducer, then you get 1 unit.
As a comparison, if you have lots of cAMP and inducer, you'd get 1000 units of B-Gal.
(If your questions come from trying a practice probelm that dealt with crp- mutants and talked about the need for cAMP in order to have maximum expression, don't worry-that question refers very specfically to materail that was taught that term).
I hope this helps
Pam
thankyou so much! i understand those
but sometimes it's very hard to differentiate "low activity" and "no activity"
my question comes from this problem:
http://img134.imageshack.us/img134/6727/topamdr1.jpg
Pam, X-gal does not turn blue in the absence of an inducer or a mutant I/O gene, right?
However, on page 721 of the textbook (right side figure), the bacteria is plated on x-gal without inducer yet it still turns blue.
Thanks
Yes, specifically engineered bacteria will turn blue even without inducer (the typical E.coli lab strains have their lac operon deleted, and partially complemented in trans by bits and pieces in plasmids, and they may or may not have the gene encoding the lac repressor, so they'll be able to turn blue).
Don't worry about it, really. A few molecules of B-Gal will be able to make some blue colour, in some cells, and the blue colour is very visible even if only a few cells in a colony make it. But I promise you that that kind of blue colour is very different frokm the nice, strong blue that you get with induction!
Yes, specifically engineered bacteria will turn blue even without inducer (the typical E.coli lab strains have their lac operon deleted, and partially complemented in trans by bits and pieces in plasmids, and they may or may not have the gene encoding the lac repressor, so they'll be able to turn blue).
Don't worry about it, really. A few molecules of B-Gal will be able to make some blue colour, in some cells, and the blue colour is very visible even if only a few cells in a colony make it. But I promise you that that kind of blue colour is very different frokm the nice, strong blue that you get with induction!
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