Research note: High efficiency transformation of the diatom Phaeodactylum tricornutum with a promoter from the diatom Cylindrotheca fusiformis

Abstract

A high efficiency transformation system was established for the pennate diatom Phaeodactylum tricornutum Bohlin using a plasmid containing fucoxanthin chlorophyll a/c binding protein (fcp) promoter/terminator and nitrate reductase (NR) promoter/terminator that are derived from the pennate diatom Cylindrotheca fusiformis. The plasmid that contains the zeocin resistance gene (ble) with the fcp promoter and enhanced green fluorescent protein gene (egfp) with the NR promoter was introduced into P. tricornutum using microparticle bombardment. Transformants (650 Ϯ 58 per 10 8 cells) were obtained. The yield of transformants was between 1.5 and 130 times higher than previously reported P. tricornutum transformation systems. Four to seven copies of the ble gene were integrated into genomic DNA of the transformants. This high efficiency transformation system of P. tricornutum is expected to provide a powerful tool for high-throughput analysis of gene function using homologous recombination or RNAi.

High-efficiency nuclear transformation of the diatom Phaeodactylum tricornutum by electroporation

“…, at least 10 times higher than reported in P. tricornutum transformation by microprojectile bombardment (Miyagawa et al, 2009;Zaslavskaia et al, 2000). Niu et al (2012) used similar electroporation parameters but a different buffer for transformation of P. tricornutum and achieved a comparable transformation efficiency to that by microprojectile bombardment.…”

“…At present, transformation of diatoms has been established for several diatom species, including Phaeodactylum tricornutum (Apt et al, 1996;Falciatore et al, 1999;Miyagawa et al, 2009;Zaslavskaia et al, 2000), Cylindrotheca fusiformis (Fischer et al, 1999;Poulsen and Kröger, 2005), Thalassiosira pseudonana (Poulsen et al, 2006), Chaetoceros sp. (Miyagawa-Yamaguchi et al, 2011), Fistulifera sp.…”

Highly Efficient Transformation of the DiatomPhaeodactylum tricornutumby Multi-Pulse Electroporation

“…3,4) Both for basic biological research and for commercial exploitation of diatoms, an efficient genetic transformation system is very important. Transformation of ditoms has been reported for several diatom species, including pennate diatoms Phaeodactylum tricornutum, [5][6][7][8] Cylindrotheca fusiformis, 9,10) and Navicula saprophila, and centric diatoms Cyclothella cryptica, 11) Thalassiosira pseudonana, 12) and Chaetoceros sp. 13) In general, delivery of foreign DNAs into diatom cells has been achieved by microprojectile bombardment, but this is not suited to high-throughput transformation of diatom cells due to the time required for sample preparation, costly consumables, and limited transformation efficiency.…”

Advances in genetic engineering of marine algae

“…Transformation methods with particle bombardment have been established for several species of diatoms, including the centric diatom Thalassiosira pseudonana (Poulsen et al, 2006), Thalassiosira weissflogii (Falciatore et al, 1999), Chaetoceros sp. (Miyagawa-Yamaguchi et al, 2011), C. cryptica (Dunahay et al, 1995), and the pinnate diatoms Navicula saprophila (Dunahay et al, 1995), P. tricornutum (Apt et al, 1996, Falciatore et al, 1999, Miyagawa et al, 2009, Zaslavskaia et al, 2000, Cylindrotheca fusiformis (Fischer et al, 1999;Poulsen and Kröger, 2005). Since 1992, biolistic methods have been applied to marine macroalgal genetic engineering (Cheney and Kurtzman, 1992).…”