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Molecular Genetics and Genomics

, Volume 265, Issue 3, pp 455–460 | Cite as

Expression of intron-containing GUS constructs is reduced due to activation of a cryptic 5′ splice site

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Original Paper

Abstract.

An intron-containing β-glucuronidase (GUS) gene has been used widely in promoter analyses and as a plant transformation marker. Maximal plant gene expression requires accurate and efficient removal of the intron from the expressed pre-mRNA transcripts by splicing. Detailed analysis of splicing of potato ST-LS1 and pea legumin introns from GUS constructs revealed the activation of a cryptic 5′ splice site in the GUS coding sequence 4 nt upstream from the authentic intron 5′ splice site. About 40% of transcripts utilised the cryptic 5′ splice site in tobacco protoplasts, reducing the translational potential of expressed pre-mRNA. The same cryptic splicing event was evident in transgenic tobacco leaves but at reduced levels. Mutations that removed the cryptic 5′ splice site are associated with a two-fold enhancement in GUS activity in tobacco protoplasts, highlighting the need for careful examination of introns and their sites of insertion into gene constructs to minimise variability in gene activity and maximise gene expression.

GUS Intron Splicing Cryptic splicing Expression levels 

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Copyright information

© Springer-Verlag 2001

Authors and Affiliations

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  1. 1.Unit of Gene Expression, Scottish Crop Research Institute, Invergowrie, Dundee DD2 5DA, UK
  2. 2.Advanced Technologies (Cambridge) Ltd., 210 Science Park, Cambridge UK

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