Syngenta patent family B

This Syngenta patent family includes patents granted in United states, Europe, Canada and Australia on  positive selection system based on  mannose or xylose. The invention of this patent family is directed to a method for selecting genetically transformed plant cells comprising the seteps of providing plant cells with a gene coding for an enzyme involved in mannose or xylose metabolism and selecting the transformed cells with mannose or its derivative or precursor. The transformed plant cells are also claimed.

Technology overview

Mannose is a hexose sugar that can strongly inhibite seed germination, root growth and respiration of plants. The sugar can be taken up by roots and converted to mannose-6-phosphate by the action of hexokinase but can not be further utilized. The accumulation of mannose-6-phosphate inhibits phosphoglucose isomerase, causing a block in glycolysis. The production of mannose-6-phosphate also depletes the cell of inorganic phosphate (orthophosphate) that is required for ATP production. While mannose has no direct adverse effect on plants, as the toxicity is not mediated by the compound itself, growth inhibition is the consequence of its phosphorylation to mannose-6-phosphate by hexokinase.

Mannose

Phosphomannose isomerase (PMI, EC 5.3.1.8) is an enzyme that converts mannose-6-phosphate to fructose-6-phosphate, an intermediate of glycolysis that positively supports the growth of plant cell.  In 1984, a gene coding for phosphomannose isomerase (manA or pmi) was first isolated from Escherichia coli by Miles and Guest.  However, its first application in plants as a selectable marker gene was reported in 1998 by Joersbo et al.  The idea was that plant cells lacking PMI are incapable of surviving on synthetic medium containing mannose as a carbon source.  Introduction of  the manA (pmi) gene into plant cells enables those transformed cells to utilize mannose as a carbon source, improve the energy status and avoid accumulation of the derivatized mannose-6-phosphate, and thus gives the transformed cell the growth advantage over the non-transformed cells.

Fructose

To date, no endogenous PMI activity has been detected in plant cells, indicating that PMI selection may be useful in the transformation of many plant species.

Another selection system is based on xylose. Some plants such as potato, tobacco and tomato can not use D-xylose but can utilize D-xylulose as the sole carbon source.  However, a problem initially encountered when xylose was used in the selection medium was the induction of callus.  This proble was solved by addition of auxin inhibitor into the selection medium.

Xylose isomerase (D-xylose ketol-isomerase, EC 5.3.1.5) catalyzes the isomerization of D-xylose to D-xylulose and the isomerization of glucose to fructose and is also termed as glucose isomerase. A gene (xylA) encoding xylose isomerase was reported to be isolated from Thermoanaerobacterium thermosulfurogenes or Streptomyces rubiginosus.

This system enables the effective selection of transgenic potato, tobacco and tomato cells using D-xylose as the selective agent.  Transgenic cells expressing the xylose isomerase gene can utilize xylose as a carbohydrate source and proliferate, whereas non-transgenic cells starve.

Specific patent information

Patent number Title, Independent Claims and Summary

Assignee

US 5767378

  • Earliest priorrity – 28 Feb 1994 (PCT)
  • Filed – 3 Oct 1995
  • Granted – 16 Jun 1998
  • Expected expiry – 28 Feb 2014
Title – Mannose or xylose based positive selection

Claim 1A method for selecting genetically transformed plant cells from a population of cells comprising:

a) introducing into plant cells a desired nucleotide sequence and a co-introduced nucleotide sequence to obtain transformed plant cells;

b) supplying to a population of plant cells including the transformed cells and non-transformed cells a compound selected from the group consisting of mannose and a derivative or precursor of mannose, wherein said transformed cells have a competitive advantage over the non-transformed plant cells in the population due to the expression or transcription of the co-introduced nucleotide sequence in the transformed cells; and

c) selecting the transformed plant cells based on said competitive advantage wherein said co-introduced nucleotide sequence is a gene encoding an enzyme involved in mannose metabolism selected from the group comprising a phosphomanno isomerase, a phosphomanno mutase, a mannose epimerase, a phosphatase, and a permease.

Claim 14Genetically transformed corn cells comprising,

an introduced desired nucleotide sequence and a co-introduced nucleotide sequence, wherein the expression or transcription of said co-introduced nucleotide sequence gives the transformed cells a competitive advantage over non-transformed cells when both transformed and non-transformed cells in a population of cells are supplied with a mannose compound and said transformed cells are selected based on said competitive advantage, wherein the co-introduced nucleotide sequence codes for a phosphosugar isomerase.

Claim 17Genetically transformed plant cells comprising an introduced nucleotide sequence and a co-introduced nucleotide sequence wherein the expression or transcription of the co-introduced nucleotide sequence gives the transformed cells a competitive advantage over non-transformed cells when both transformed and non-transformed cells in a population of cells are supplied with a compound and said transformed cells are selected based on said competitive advantage, wherein the co-introduced nucleotide sequence is a gene encoding an enzyme involved in mannose metabolism selected from the group comprising a phosphomanno isomerase, a phosphomanno mutase, a mannose epimerase, a phosphatase, and a permease.

Originally assigned to Novartis AG, and then reassigned to Syngenta Participations AG

EP 804599 B1

  • Earliest priorrity – 28 Feb 1994
  • Filed – 5 Nov 1997
  • Granted – 24 May 2006
  • Expected expiry – 5 Nov 2017
Title – Mannose or xylose based positive selection

Claim 1
A method for identifying or selecting from a population of plant cells selected from the group consisting of fruits, small grain cereals, vegetables, canola, sunflower, tobacco, sugar beet corn and cotton cultivated or in a medium containing at least one compound, cells which have a metabolic advantage as a result of having being transformed, wherein:i) the cells are transformed with a nucleotide sequence or a co-introduced nucleotide sequence one of which comprises a region which encodes an enzyme protein which is involved in the metabolism of the compound selected from the group consisting of phosphosugar-isomerases, phosphosugar-mutases, phosphatases, and sugar-epimerases;ii) the compound is mannose or xylose or a derivative or a precursor of these, or a substrate of the enzyme protein involved, either directly or indirectly, in the metabolism of mannose or xylose.

Syngenta Participations AG

AU 682495 B2

  • Earliest priorrity – 28 Feb 1994
  • Filed – 28 Feb 1994
  • Granted – 9 Oct 1997
  • Expected expiry – 28 Feb 2014
Title – Mannose or xylose based positive selection

There is no specification available online for this patent, Should we get the PDF from IPaustralia?

Originally assigned to Sandoz Ltd, and then reassigned to Syngenta Participations AG

Remarks

Related patent application was filed in Canada (CA 2157470). Patents were also granted in Japan (JP 3698722 B2) ans Russia (RU 2126834). A PCT application (WO 94/20627) was also filed.

Search strategy

Search details
Date of search 25/05/2006
Database searched Patent Lens
Type of search Simple, stemming on
Collections searched AU-B, US-A, US-B, EP-B, WO
Search terms “Novartis” in applicant

“mannose” in abstract

Results 3 hits
Comments Only US 5767378 is the relevant patent on positive selection