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Terminator sequences for plant gene expression

一种转录终止子、序列的技术，应用在植物分子生物学和植物基因工程领域

Inactive Publication Date: 2017-10-13
PIONEER HI BRED INT INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Read-through transcription through a gene's leaky terminator can lead to unwanted transcription of one transgene from another gene's promoter

Method used

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  • Terminator sequences for plant gene expression
  • Terminator sequences for plant gene expression
  • Terminator sequences for plant gene expression

Examples

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example

[0106] The invention will be further illustrated in the following examples, wherein parts and percentages are by weight and degrees are degrees Celsius unless otherwise stated. It should be understood, that these Examples, while indicating embodiments of the invention, are given by way of illustration only. From the above discussion and these examples, one skilled in the art can ascertain the essential characteristics of this invention, and without departing from the spirit and scope thereof, can make various changes and modifications to the invention so as to be applicable to various usage and conditions. Furthermore, various modifications of the invention in addition to those shown and described herein will be apparent to those skilled in the art from the foregoing description. Such modifications are also intended to be within the scope of the appended claims.

example 1

[0108] Amplification and Cloning of Sorghum bicolor γ-kafirin Terminator Sequence

[0109] Primers (SEQ ID NOS: 2 and 3) were designed according to the Sorghum bicolor genome sequence database for amplifying the terminator of the γ-kafirin gene (SB-GKAF) from Sorghum bicolor. The primer sequences are given below, the underlined regions are nonlogous to the genomic template:

[0110] TMS2039 (forward primer; SEQ ID NO: 2):

[0111] CAGATCTGATATCGATGGGCCCACT AACTATCTATACTGTAATAATGTTGTATAG

[0112] TMS2040 (reverse primer; SEQ ID NO: 3):

[0113] CGGACCGGGTGACC AAGCTTAAGCGAACATATGTCCCTC

[0114] Using these primers, a 504 bp product containing the 465 bp SB-GKAF terminator sequence (SEQ ID NO: 1 ) was amplified by PCR. The product was cloned into pGEMTeasy (Promega) (PHP31801; figure 1 ; SEQ ID NO: 4) and confirm the sequence. The cloned SB-GKAF terminator included 165 bp of the predicted 3' untranslated region (UTR) of SB-GKAF and about 300 bp of downstream sequence...

example 2

[0119] Transient transformation to test the efficacy of the SB-GKAF terminator

[0120] The ability of the isolated SB-GKAF terminator sequence (SEQ ID NO: 1 ) to function efficiently as a terminator was tested in recombinant constructs. Its efficacy as a terminator was tested by its ability to terminate transcription and by its ability to increase protein expression. Since incorrect terminators can lead to incorrect processing of the 3' end of the mRNA, thereby affecting RNA stability, terminators have been found to affect protein expression levels. It has been shown that different terminators can lead to up to 100-fold variation in transgene expression efficiency (Bieri et al., (2002) Molecular Breeding 10: 107-117; An et al. (1989) Plant Cell 1: 115-122; Ingelbrecht et al. (1989) , Plant Cell, 1:671-680; Ali and Taylor (2001) Plant Mol. Bio., 46:251-261). We thus tested the ability of the SB-GKAF sequence (SEQ ID NO: 1 ) to increase protein expression compared to the we...

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Abstract

The invention discloses polynucleotide sequences that can be used to regulate gene expression in plants. A plant functional terminator sequence derived from Sorghum bicolor is disclosed.

Description

[0001] Cross references to related patent applications [0002] This patent application claims the benefit of US Provisional Application No. 61 / 514,055, filed August 2, 2011, which is hereby incorporated by reference in its entirety. technical field [0003] The invention relates to the fields of plant molecular biology and plant genetic engineering. More specifically, it relates to novel plant terminator sequences and their applications for regulating plant gene expression. Background technique [0004] Recent advances in plant genetic engineering have opened new doors for engineering plants to have improved traits or traits. These transgenic plants typically have in their genome a recombinant DNA construct having a protein coding region operably linked to regulatory regions that allow for precise expression of the transgene. Some examples of regulatory elements that help regulate gene expression in transgenic plants are promoters, introns, terminators, enhancers and sil...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/82A01H5/00
CPCC12N15/8216C12N15/00
Inventor S.E.阿比特R.容
Owner PIONEER HI BRED INT INC
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