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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

  1. RE3 (0.25 marks)

  2. 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)

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