User:Paulina Alatriste/Notebook/UNAM Genomics Mexico 2011/2011/08/09

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Planning of the assembly of parts/Oligo design

ABSTRACT

  • First planning of the assembly of parts in order to present it at the team meeting.
  • Oligo design

  • The synthesis is almost here, so we need to have an action plan to be efficient and take advantage of the time we have. So today Fibo and I started to plan some aspects of the assembly of the parts, this are our conclusions:
-The parts come in six plasmids, we need to obtain all the fragments with PCR using oligos that have an overlapping region with other parts in order to perform Gibson assembly.
-In a first step of ligation the parts that where synthesized separated into halves are going to be assembled. This is the case of PFOR and HydEF where each halve of the gene comes in different plasmids.
-In a second step of ligation PFOR-HydA and HydEF-HydG are going to be assembled.
-We have to design and order the oligonucleotides immediately to amplify the regions out of the plasmids. It is necessary to do this design consciously, because they need to have determined overlapping regions. 
-We need to send to the registry all the parts separated, in total 4 parts.
-We need to find out what is the best: clean the PCR product and then isolate it from a gel band, or only isolate the product from a gel band.
  • Today we have team meeting so we are going to present our conclusions to all the team members. We expect a discussion and feedback.

  • We accorded in the team meeting that 8 pairs of oligos have to be made, so we assigned one pair to one team member and that team member has to design the oligos.



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