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The bacterial communities associated with microalgae are vital for the growth and health of the host, and engineering algal microbiomes can enhance the fitness of the algae. Characterization of these microbiomes mostly relies on sequencing of DNA, which can be extracted with an array of protocols that potentially impact DNA quantity and quality and thus potentially affect subsequent analyses of microbiome composition. Here, we extracted DNA from Isochrysis galbana, Tetraselmis suecica, and Conticribra weissflogii microbiomes using four different protocols. DNA yield and quality was greatly impacted by the choice of extraction protocol, whereas microbiome composition determined by 16S rRNA gene amplicon sequencing was only impacted to a minor degree, with microalgal host species being the main determinant of microbiome composition. The I. galbana microbiome was dominated by the genus Alteromonas , whereas the microbiome associated with T. suecica was dominated by Marinobacteraceae and Rhodobacteraceae family members. While these two families were also prevalent in the microbiome associated with C. weissflogii , Flavobacteriaceae and Cryomorphaceae were also highly dominant. Phenol-chloroform extraction resulted in higher DNA quality and quantity compared to commercial kits; however, because they have other advantages such as high throughput and low toxicity, commercial kits can be employed to great benefit for the characterization of microalgal microbiomes. IMPORTANCE Microalgae are very important as primary producers in the ocean, but also as forthcoming sustainable producers of biotechnologically interesting compounds. Accordingly, the bacterial microbiomes associated with microalgae are attracting increasing attention due to their effects on the growth and health of microalgae. Since most members of these microbiomes cannot be cultured, knowledge about community composition is best obtained using sequencing-based methods. This study evaluates the impact of DNA extraction methods on DNA quantity and quality along with sequence-based characterization of the bacterial microbiome composition of three microalgae: Isochrysis galbana, Tetraselmis suecica, and Conticribra weissflogii.