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medium choice confusion

Discussion of all aspects of biological molecules, biochemical processes and laboratory procedures in the field.

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medium choice confusion

Postby ashhh » Sun Aug 21, 2011 2:58 pm

Hi everyone, this is my first post here. I was looking through past papers and came across my nemesis- bacterial growth medium. Heres the question:

Q. You have been provided with the following E.coli strains to use in conjugation mapping (interrupted mating) experiments in order to construct a genetic map.

Hfr strains
Hfr 1 str s (sensitive to streptomycin)
Hfr 2 azi s (sensitive to sodium azide)

The Hfr strains are wild type for all the amino acid and essential nutrient genetic markers and are sensitive to streptomycin or sodium azide as indicated.

F- strain
LAN100 F- thi -, trp -, his -, met -, arg -, thr -, phe -, pro-, str r, azi r

These mutations confer the following phenotypes on the strains:
Genotype Phenotype
thi - Thi- cannot synthesise vitamin B1 (thiamine)
trp - Trp- cannot synthesise tryptophan
his - His- cannot synthesise histidine
met - Met- cannot synthesise methionine
arg - Arg- cannot synthesise arginine
thr - Thr- cannot synthesise threonine
phe - Phe- cannot synthesise phenylalanine
pro- Pro- cannot synthesise proline
str r Str r resistant to streptomycin
azi r Azi r resistant to sodium azide

a)What is the simplest defined medium you can use to grow the F- strain LAN100?

b) What medium would you use to select His+ exconjugants in a mating experiment between Hfr1 and strain LAN100?

c) What medium would you use to select Arg+ exconjugants in a conjugation mapping experiment between Hfr2 and your LAN100 F- strain?

I know these are really simple but I'm never confident. My guess is that a) would consist of the + of each genotype? whereas b) and c) are ... all but his+ and arg+ respectively?

Thanks for any guidance!
ashhh
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Postby canalon » Mon Aug 22, 2011 9:36 am

for a) you need to provide all the missing things that the stairn cannot provide by itself.
for b) and c) you want to provide everything but the missing gene, so only those that have acquired the gene will grow, but you need to select against the donor too, so you need to use an antimicrobial to kill it too (ie str or azide)
Patrick

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