Part:BBa_K2797002

Composite part for streptomycin resistance

This composite part confers resistance to streptomycin. The part contains the constitutive promoter BBa_J23119 from the Anderson collection, BBa_B0034 ribosome binding site, BBa_K125520 coding sequence conferring resistance to streptomycin and BBa_K2797001 terminator.

Usage and Biology

A range of antibiotic resistance genes are required for selection of transformants when inserting foreign DNA into a chassis organism. Newcastle iGEM 2018 focused on developing a new chassis organism capable of colonising plant roots. The selected organism, Pseudomonas sp. DSM 25356 was found to be resistant to a range of commonly used antibiotics. Streptomycin was one of two antibiotics Pseudomonas sp. was found not to be resistant to. The team has deposited a new streptomycin resistance cassette.

Characterisation

Figure 1 E. coli strain DH5α transformed with streptomycin resistance part BBa_K2797002 plated on LB agar containing streptomycin (50 μg/ml).

Figure 2 E. coli strain DH5α plated on LB agar containing streptomycin (50 μg/ml).

Figure 3 E. coli strain DH5α carrying the streptomycin resistance part BBa_K2797002 plated on LB agar containing chloramphenicol (25 μg/ml). The part is in the pSB1C3 backbone conferring resistance to chloramphenicol.

Figure 4 E. coli DH5α with or without the BBa_K2797002 part in pSB1C3 were grown in LB medium containing streptomycin at varying concentrations. Cells were grown in 96-well plate format in 200 μl volumes at

37&#176 C

over 24 hours. (n=3 replicates, error bars are standard error of the mean).

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 7
  Illegal NheI site found at 30
  Illegal NheI site found at 861
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 709
  
• 1000
  COMPATIBLE WITH RFC[1000]