While working in Dr. Strap’s lab you identify a number of E. coli mutants that o
ID: 218607 • Letter: W
Question
While working in Dr. Strap’s lab you identify a number of E. coli mutants that only grow in the presence of cysteine. Your supervisor suggests that the next step is for you to map the cys gene you have discovered relative to other known mutations. To do this, you start with bacterial strains:
Hfr cys- gal+ his+ lac+ pur+ trp+ tons
F- cys+ gal- his- lac- pur- trp- tonR
cys- his- pur- trp- = auxotrophic for cysteine, histidine, purines, tryptophan.
lac- gal- = cannot metabolize lactose or galactose.
tons = sensitive to phage T1.
a. If you want to select for cys+, lac+ and tonR recombinants resulting from a cross between the Hfr and F- strains, what media would you need to plate the bacteria on? That is, what nutrients would the minimal media need to be supplemented with?
b. If you supplement the minimal media with glucose, histidine, purines and T1 phage, what recombinant classes will you grow?
Explanation / Answer
Ans .
a) to select for cys+, lac+ and tonR recombinants resulting from a cross between the Hfr and F- strains, the minimal media should be supplemented with galactose as a carbon source instead of glucose (to select for lac+ recombinants i..e.,only bacteria which are capable of metabolizing lactose will grow) and T1 phages (to select for tonR recombinants i.e., only bacteria which are resistant to T1 will grow ).
to select for cys+ recombinants the minimal media should not contain cysteine .
b) in the minimal media with glucose, histidine, purines and T1 phage, following recombinant classes will grow
cys+ gal- his- lac- pur- trp+ tonR
cys+ gal+ his- lac- pur- trp+ tonR
cys+ gal+ his+ lac- pur- trp+ tonR
cys+ gal+ his+ lac+ pur- trp+ tonR
cys+ gal+ his+ lac+ pur+ trp+ tonR
both gal- and gal+ (and lac+ and lac- )will be able to grow on minimal media supplemented with glucose . this is because glucose is the preferred carbon source and as long as glucose is present in the medium, the ability to metabolize lactose or galactose doesn't make any differance.
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