Part:BBa_K3930004

Up integrative sequence to target locus XII-1 of S. cerevisiae genome Sequence and Features

Assembly Compatibility:

10
INCOMPATIBLE WITH RFC[10]
Illegal PstI site found at 256
12
INCOMPATIBLE WITH RFC[12]
Illegal PstI site found at 256
21
INCOMPATIBLE WITH RFC[21]
Illegal BglII site found at 293
23
INCOMPATIBLE WITH RFC[23]
Illegal PstI site found at 256
25
INCOMPATIBLE WITH RFC[25]
Illegal PstI site found at 256
1000
COMPATIBLE WITH RFC[1000]

Introduction

The sequences to target the integration locus XII-1 (Chr XII: 795787..796720) of S. cerevisiae genome come from the EasyClone-MarkerFree kit (Jessop-Fabre et al., 2016). This part is flanking the insert in 5' and must be used with the (BBa_K3930005) XII-1 down part in 3'.

Results

Integration of the part (BBa_K3930000) into the yeast genome locus XII-1

The part (BBa_K3930000) was linearized and transformed into the S.cerevisiae LycoYeast strain according to the Amandine transformation protocol, with 2 µg of DNA. The construction is flanked by parts (BBa_K3930004) and (BBa_K3930005). Figure 1 shows the electrophoresis gel of PCR on colony to check the clones (clone 3 is OK).

Figure 1: Integration of pFLEUR in LycoYeast at locus XII-1

pFLEUR was checked with agarose electrophoresis gel and revealed with EtBr. A theoretical gel is presented on the right of each gel (note that a different ladder is presented on the theoretical gel)

The Integrative locus XII-1 up (BBa_K3930004) coupled with the integrative locus (BBa_K3930005) XII-1 down part is functional to target integration at the XII-1 locus under those experimental conditions.

References

Chen X, Shukal S, Zhang C. 2019. Integrating Enzyme and Metabolic Engineering Tools for Enhanced α-Ionone Production. J Agric Food Chem. 67(49):13451–13459. doi:10.1021/acs.jafc.9b00860.