Difference between revisions of "Part:BBa K3909018"

Revision as of 10:36, 16 October 2021

pYLXP-ylPOT1-ylMFE1-ylPOX1

This composite part consist of pYLXP’-ylPOT1 (BBa_K3909010), pYLXP’-ylMFE1 (BBa_K3909011) and pYLXP’-ylPOX1 (BBa_K3909012). As shown in figure 1, our biobricks design is mainly divided into two parts: fatty acid degradation and γ-linolenic acid synthesis. The BBa_K3909018 is belong to part one --fatty acid degradation.

Fig.1 The overview of our biobricks design.

We plan to enhance the oli degradation pathway by expressing three endogenous fatty acid degradation genes ylMEF1 (BBa_K3909006), ylPOT1 (BBa_K3909007), and ylPOXn (from BBa_K3909000 to BBa_K3909005), which are related to the metabolim of transforming acyl-CoA into acetyl-CoA in peroxisome (β-oxidation). Specifically, the β-oxidation includes three steps: i) oxidation, that catalyzed by six acyl-CoA oxidases (translated from ylPOX1 to ylPOX6); ii) hydration and dehydration, that catalyzed by multifunctional enzyme (translated from ylMFE1); and iii) thiolysis, that catalyzed by 3-ketoacyl-CoA thiolase (translated from ylPOT1)[1]. Firstly,we constructed the single gene-overexpressed plasmids by the aforementioned method, including pYLXP’-ylPOT1(BBa_K3909010), pYLXP’-ylMEF1 (BBa_K3909011) and pYLXP’-ylPOXn (from BBa_K3909012 to BBa_K3909017) , which are shown in Figure 2.

Fig.2 Single fragment plasmid construction.

For overpressing the whole oil degradation pathway, we further combined these genes by BioBrick method, and constructed plasmids pYXLP’-ylPOT1-ylMFE1-ylPOXn (from BBa_K3909018 to BBa_K3909023), as shown in figure 3. The Y. lipolytica (Po1f) we used cannot synthesize leucine by itself, so the Leu2 marker on the plasmid can be used as marker gene. Then, the constructed plasmids were transformed into Y. lipolytica Po1f.

Fig.3 Multi-fragment plasmid construction.

Sequence and Features