Team:IISER-Pune-India/TeamNotebook/Team Notebook 11e9906b405543a29361e38a50c81024/Questions 1 - Wet Lab 2866b6ef8f6342f8b0f12780fa088b4f

Questions 1 - Wet Lab

Questions 1 - Wet Lab

DepartmentWet Lab
DescriptionList of questions we need to find answers to
HP sub-branch
Property 1
Property 2

Note: Please put up annotated/highlighted PDFs of papers in the 'Answers' pages

After General Wet Lab Meeting 1 , we came up with a set of questions to be answered.

They are as follows:

General Questions

  • What are the list of parts we will need for the planned genetic modifications?
  • What are the corresponding BioBricks for these parts if any?
  • What are the methods we will use to perform these modifications? (contact Prem and PhD mentors accordingly)
  • What data can we get from Dry Lab that makes our life easier?

Please note that these questions have to be answered by all 3 groups

Questions specific to E. coli

  • What strain of E. coli are we using?
  • What is the final metabolic pathway expected to be after all engineering is done?
    • Do we stick exactly to the Lin et al 2005 paper?
  • How are knockouts done? (contact PhD mentors)
  • What promoters are we going to put the genes under? (contact PhD mentors)
  • What will be the final construct of the plasmid we will be using? (contact PhD mentors)
  • How do we get the succinate out of the E. coli?
  • How efficient should sucrose uptake be so as to not overwhelm the cyanobacteria?
  • What is the final list of modifications that need to be made?
  • What genes are we editing?
    • glgc/invertase v/s sps/spp
    • What are the final constructs we will be using for the transformation? (contact Prem)

Questions Specific to Cyanobacteria

  • What strain of cyanobacteria are we using?
  • What promoters are we going to put the genes under?
  • What is the method of transformation that is most preferred for the strain we choose? (contact Prem)

Questions Specific to the co-culture

  • How to make similar the growth rates of cyanobacteria and E. coli?
    • Media?
    • Temperature?
  • Do we encapsulate the cyanobacteria and E. coli in the co-culture?
    • Does this make assaying the growth rate easier?
  • How do we get succinate out of the growth medium?
  • What is the optimum salt stress & pH for the co-culture? + how will we maintain constancy in levels
  • Assays:
    • Sucrose production
    • Succinate production
    • Rate transfer assays for sucrose
    • Expression assays (Wild Type v/s Mutant)