Part:BBa_K4879005

Yarrowia lipolytica ScRAD52 gene

RAD52 coding sequence from Saccharomyces cerevisiae, codon optimized for Yarrowia lipolytica. [1]

Usage and Biology

RAD52 is a well-documented protein that helps in facilitating homologous recombination and subsequent DNA repair; found to be well-conserved across many species. It assists in the formation of a nucleoprotein filament on the broken single-stranded DNA (ssDNA) and thus facilitates strand invasion, the first step in homologous DNA repair.

Wild-type Y.lipolytica has a very low probability of homologous recombination, which was detrimental to the deletion attempts we made to delete faa1 and alk2 genes from our chassis. As ScRAD52 enhances this probability to as high as 90%, we inserted the (ScRAD52 expression construct) into our organism.

The protein structure of ScRAD52, as predicted by AlphaFold.

Design

The transcriptional unit for CvFAP (BBa_K4879011) is designed with the coding sequence flanked by the TEF1 promoter (BBa_K2983053) upstream and the XPR2 terminator (BBa_K3629004) downstream.

References

1. Qingchun Ji, Jie Mai, Ying Ding, Yongjun Wei, Rodrigo Ledesma-Amaro, Xiao-Jun Ji, Improving the homologous recombination efficiency of Yarrowia lipolytica by grafting heterologous component from Saccharomyces cerevisiae, Metabolic Engineering Communications, Volume 11, 2020, e00152, ISSN 2214-0301, https://doi.org/10.1016/j.mec.2020.e00152.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 403
  Illegal BglII site found at 533
  Illegal XhoI site found at 1223
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 1207
  
• 1000
  COMPATIBLE WITH RFC[1000]

References

1. Qingchun Ji, Jie Mai, Ying Ding, Yongjun Wei, Rodrigo Ledesma-Amaro, Xiao-Jun Ji. Improving the homologous recombination efficiency of Yarrowia lipolytica by grafting heterologous component from Saccharomyces cerevisiae, Metabolic Engineering Communications, Volume 11, 2020, e00152, ISSN 2214-0301 https://doi.org/10.1016/j.mec.2020.e00152