Gene transfer of brine shrimp (Artemia sinica) by electroporation

• Main Authors: Yen-Ta Chen, 陳彥達
• Other Authors: Ying-Chou Lee
• Format: Others
• Language: zh-TW
• Published: 2006
• Online Access: http://ndltd.ncl.edu.tw/handle/31547177449926547359

碩士 === 國立臺灣大學 === 漁業科學研究所 === 94 === The nauplii of crustacean Artemia, also named brine shrimp, are the most widely used live zooplankton bait for feeding larvae of aquaculture finfish and shellfish. By taking advantages of genetically transformed Artemia, some traits of the feeder can be improved indirectly such as fast-growing or disease-resistance. This approach might decrease the potential risk of using genetically modified finfish and shellfish. Here we developed a gene transfer method which enabled to introduce exogenous DNA into the genome of target species－Artemia sinica efficiently. Different DNA constructs were introduced into the decapsulated cysts of Artemia by electroporation under 2KV, 20 μsec pulse length, 20 number of pulses and a DNA concentration of 20μg/μl in a volume of 100μl. Among seven different constructions we examined, a DNA fragment, named pCMV-MB-EGFPITR, containing EGFP reporter gene driven by human cytomegalovirus (CMV) promoter and medaka β-actin promoter, produced the most intense GFP expression. Although the 24 hour-hatching rate of Artemia nauplii (30±1％) was significantly lower than untreated group (61±7％), the GFP expression rate was 33±6％ in 72 hours after hatching. Genomic DNA was extracted from the F0, F1 and F2 generation of transgenic Artemia, a 719-bp product was amplified which was correspondent with the reporter gene as a positive control, by polymerase chain reaction. These evidences suggest that transgene was integrated into the genome of transgenic Artemia and inherited to their offsprings. This is the first report which demonstrates that a transgenic line of Artemia with strong fluorescent expression can be generated.