And the accuracy and efficiency translation
The binding specificity in this interaction can be influenced by specific modifications that occur in the anticodon region of the tRNA. One such modification requires a functional methylaminomethyl (mnm) pathway (Figure 1). This pathway modifies the N34 position of the anticodon with a methylaminomethyl group which subsequently can affect codon recognition, and the accuracy and efficiency of translation.
PCR amplified upstream and downstream regions of the mnmE gene
A cassette containing a gusA reporter gene and the nptII gene encoding kanamycin resistance
*Cm, Chloramphenicol; Nx, Naladixic acid, Sm, Streptomycin; Sp, Spectinomycin; Tm, Trimethoprim
| Plasmid | Characteristic1 | Antibiotic resistance to2 |
|---|---|---|
| Plasmid A | Mobilisable BHR plasmid | Km |
| Plasmid B | Mobilisable NHR helper plasmid | Cm |
| Plasmid C | Non mobilisable BHR plasmid | Gm |
| Plasmid D | Mobilisable NHR plasmid | Km |
| Plasmid E | Mobilisable BHR plasmid | Gm |
| Plasmid F | Mobilisable NHR plasmid | Gm |
Figure 1: Maps of Plasmids available to you. Consult Table 2 as well.
Gene A = SMED_RS16080
tRNA uridine-5-carboxymethylaminomethyl(34) synthesis enzyme MnmG
Q4 Which plasmid in Figure 1 and Table 2 would you choose as the backbone to create a suicidal QL inactivation plasmid for use with E. medicae WSM419? (0.5 mark)
NotI 9.8
XhoI 9.8
XhoI and EcoRI 6.2, 2.7, 0.9
NotI and EcoRI 3.6, 3.6, 2.6
RE3 (0.25 marks)
RE4 (0.25 marks)
Q8 Using the diagram below, fill in the following Table to give the names of the genes that are present in the mnmE gene neighbourhood following single crossover homologous recombination. (1.5 marks)
| Letter for gene in diagram | Gene name |
|---|---|
| A | |
| B | |
| C | |
| D | |
| E | |
| F |
Q12 What free-living phenotype might you expect for a mnmE E. medicae mutant? (1.0 marks)
Total marks (15)