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Plant Biotechnol J. 2010 Sep;8(7):772-82. doi: 10.1111/j.1467-7652.2010.00504.x. Epub 2010 Mar 16.

Revealing frequent alternative polyadenylation and widespread low-level transcription read-through of novel plant transcription terminators.

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1
DuPont/Pioneer Crop Genetics, Experimental Station, Wilmington, DE, USA.

Abstract

Plant genetic engineering can create transgenic crops with improved characteristics by introducing trait genes through transformation. Appropriate regulatory elements such as promoters and terminators have to be present in certain configurations for the transgenes to be properly expressed. Five terminators native to soybean genes-encoding a MYB family transcription factor (MYB2), a Kunitz trypsin inhibitor (KTI1), a plasma membrane intrinsic protein (PIP1), a translation elongation factor (EF1A2) and a metallothionein protein (MTH1) were cloned and tested for their ability to enable transgene expression, mRNA polyadenylation and transcription termination. The terminators are as good as a control terminator of the potato proteinase inhibitor II gene (PINII) in conferring proper transgene expression, leading to mRNAs with various polyadenylation sites and terminating mRNA transcripts. RNA transcription read-through was detected in all transgenic plants and was quantified by qRT-PCR to be <1% at positions approximately 1 kb downstream of the 5' ends of different terminators. The detection of read-through RNA transcripts of the corresponding endogenous genes up to approximately 1 kb beyond the polyadenylation sites suggests that limited RNA transcription read-through is a normal phenomenon of gene expression. The study also provided more choices of terminators for plant genetic engineering when constructing DNA constructs containing multiple gene expression cassettes.

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