TY - JOUR
T1 - Enhancement of efficient molecular biology tools for Tremella fuciformis spore
AU - Yu, Caiyuan
AU - Sun, Yuanwei
AU - Yang, Chunkai
AU - Fang, Yan
AU - Meng, Qi
AU - Qiu, Yibin
AU - Lei, Peng
AU - Xu, Hong
AU - Zhang, Qi
AU - Li, Sha
N1 - Publisher Copyright:
© 2024 Elsevier Ltd
PY - 2024/8
Y1 - 2024/8
N2 - Tremella fuciformis, one of the most approved mushrooms worldwide, possesses both edible and medicinal value. However, low transformation efficiency and high false positive rate, long verification period, and low expression of heterologous proteins hinder further genetic research. In this study, a polyethylene glycol-mediated genetic transformation system was established by using the homologous resistance marker carboxin in T. fuciformis. By optimizing the protoplast quality and transformation conditions, the transformation efficiency reached 8.23 CFU/μg, and the positive rate reached 64.95%. Moreover, Driselase-Heating-Freezing screening system was developed for rapid transformants verification, which took only 35–45 min and made the colony PCR 100% accurate. Considering the rationality of tRNA use frequency in heterologous genes, the coordinated relative synonymous codon usage (RSCU) strategy was adopted to recode the heterologous gene sequence, which ensured the protein expression level. This study comprehensively enhances molecular biology tools in T. fuciformis, and lays a foundation for further development.
AB - Tremella fuciformis, one of the most approved mushrooms worldwide, possesses both edible and medicinal value. However, low transformation efficiency and high false positive rate, long verification period, and low expression of heterologous proteins hinder further genetic research. In this study, a polyethylene glycol-mediated genetic transformation system was established by using the homologous resistance marker carboxin in T. fuciformis. By optimizing the protoplast quality and transformation conditions, the transformation efficiency reached 8.23 CFU/μg, and the positive rate reached 64.95%. Moreover, Driselase-Heating-Freezing screening system was developed for rapid transformants verification, which took only 35–45 min and made the colony PCR 100% accurate. Considering the rationality of tRNA use frequency in heterologous genes, the coordinated relative synonymous codon usage (RSCU) strategy was adopted to recode the heterologous gene sequence, which ensured the protein expression level. This study comprehensively enhances molecular biology tools in T. fuciformis, and lays a foundation for further development.
KW - Codon usage preference
KW - Genetic transformation system
KW - Heterologous gene expression
KW - Homologous resistance marker
KW - Tremella fuciformis spore
UR - http://www.scopus.com/inward/record.url?scp=85194064708&partnerID=8YFLogxK
U2 - 10.1016/j.fbio.2024.104399
DO - 10.1016/j.fbio.2024.104399
M3 - 文章
AN - SCOPUS:85194064708
SN - 2212-4292
VL - 60
JO - Food Bioscience
JF - Food Bioscience
M1 - 104399
ER -