Chromosomal integration of the Vitreoscilla hemoglobin gene and its physiological actions in Tremella fuciformis.
The Vitreoscilla hemoglobin (VHb) gene was expressed in yeast-like conidia (YLCs) of Tremella fuciformis (T. fuciformis) to increase cell density in submerged fermentation by enhancing oxygen uptake. With the intention of doing this, an integrated expression vector containing the VHb gene and the hygromycin B phosphotransferase (hph) gene derived from Escherichia coli (E. coli) as the selectable marker was constructed, and then transformed into protoplasts of YLCs from T. fuciformis with restriction enzyme-mediated DNA integration (REMI). Hygromycin-resistant transformants had been generated during the transformation. Molecular evidences including PCR assay, Southern blotting, and Western blot analysis indicated the VHb gene had been integrated into the genome of transgenic T. fuciformis strains and was expressed successfully. Shake-flask fermentation and bioreactor cultivation results showed that the expression of VHb in this fungus could enhance growth of YLCs. The final cell density was higher in the culture of VHb-expressing strain than that of the wild-type strain. Moreover, these results also suggested that CaMV35S promoter was capable of driving the expression of heterologous genes in T. fuciformis.
Zhu
Hu
H
State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai 200237, People's Republic of China.
Wang
Tian-Wen
TW
Sun
Shu-Jing
SJ
Shen
Ya-Ling
YL
Wei
Dong-Zhi
DZ
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D001487:Basidiomycota / Q000378:metabolism*; D006454:Hemoglobins / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D010101:Oxygen Consumption / Q000502:physiology; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D054793:Truncated Hemoglobins; D020589:Vitreoscilla / Q000235:genetics*
D001426:Bacterial Proteins; D006454:Hemoglobins; D011993:Recombinant Fusion Proteins; D054793:Truncated Hemoglobins; C082613:hemoglobin protein, Vitreoscilla