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Sample Preparation and Analysis of RNASeq-based Gene Expression Data from Zebrafish
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Increased functional protein expression using nucleotide sequence features enriched in highly expressed genes in

Eric J Horstick1, Diana C Jordan1, Sadie A Bergeron1

  • 1Program in Genomics of Differentiation, Eunice Kennedy Shriver National Institute of Child Health and Human Development, Bethesda, MD 20892, USA.

Nucleic Acids Research
|January 29, 2015
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Summary

Scientists engineered zebrafish gene expression by optimizing sequence features. This biomolecular engineering approach significantly boosts protein yield in zebrafish, offering a method for other animal models.

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Area of Science:

  • Molecular Biology
  • Genetics
  • Biotechnology

Background:

  • Protein expression levels are crucial for genetic manipulations but are often limited.
  • Identifying and assessing the impact of specific nucleotide sequence features on protein expression is challenging.

Purpose of the Study:

  • To identify and validate sequence features that enhance protein expression in zebrafish embryos.
  • To develop principles for biomolecular engineering to increase protein yield in zebrafish.

Main Methods:

  • Analyzed RNAseq and microarray data to find highly expressed genes in zebrafish embryos.
  • Systematically analyzed sequence features correlated with protein expression levels.
  • Tested identified features using mRNA microinjection and functional protein reporter assays in zebrafish embryos.

Main Results:

  • Codon selection, releasing factor recognition sequences, and specific introns/3' untranslated regions were found to increase protein expression.
  • Protein expression was enhanced 1.5- to 3-fold by these identified features.
  • Developed software (CodonZ) and plasmid vectors based on these principles for rational gene design.

Conclusions:

  • Rational gene design principles significantly improve protein expression in zebrafish.
  • The identified sequence features and design approach can be applied to enhance protein yield in other animal models.