# US 9,872,504 B2 · Bread yeast strains

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- Number: US 9,872,504 B2
- Document ID: US9872504B2
- Status: Granted
- Kind: B2
- Application number: 13/922,634
- Filed: 2013-06-20
- Published: 2013-12-26
- Granted: 2018-01-23
- Earliest claimed: 2010-12-21
- Assignees: Lesaffre Et Compagnie
- Applicants: Lesaffre Et Compagnie
- Inventors: Jean-Charles Bartolucci; Didier Colavizza; Melanie Legros; Georges Pignede
- Web page: https://patentgrep.com/patent/US9872504B2

## CPC

- A21D 8/047: {with yeasts}
- C12N 1/18: Baker's yeast; Brewer's yeast
- C12N 9/2408: {acting on alpha -1,4-glucosidic bonds}
- C12N 9/2431: {Beta-fructofuranosidase (3.2.1.26), i.e. invertase}
- C12N 15/81: for yeasts
- C12Y 302/0102: Alpha-glucosidase (3.2.1.20)
- C12Y 302/01026: Beta-fructofuranosidase (3.2.1.26), i.e. invertase
- A21D: TREATMENT OF FLOUR OR DOUGH FOR BAKING, e.g. BY ADDITION OF MATERIALS; BAKING; BAKERY PRODUCTS (preservation of flour or dough before baking A23B40/00; preservation of bakery products A23B45/00)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- C12Y: ENZYMES

## Abstract

The present invention relates to novel broad spectrum strains of baker's yeast, i.e. yeast strains that are effective on both dough known as unsweetened dough and dough known as sweet dough.

## Claims

### Claim 1

A method for obtaining a broad spectrum strain of Saccharomyces cerevisiae, comprising the steps of:
selecting a strain of Saccharomyces cerevisiae having:
an invertase activity of 300 units or more;
a maltase activity of less than 80 units; and
a fermentative capacity of 110 mL or more in an unsweetened dough; and
inactivating m copies of a SUC gene of said selected strain by disruption, m being a whole number equal to 10, 11 or 12, such that the invertase activity of the selected and inactivated strain is 20 units or less, wherein the inactivation step comprises:
a step a) of transforming said selected strain with a disruption cassette comprising a selection marker flanked by two lox sequences, said lox sequences being flanked by two homologous recombination sequences, wherein each of said two homologous recombination sequences comprises a fragment of SEQ ID NO: 3;
a step b) of selecting transformed strains that have integrated the disruption cassette; and
an optional step c) of eliminating the selection marker,
wherein the ratio between invertase and maltase is altered from at least 5:1 before inactivation to at least 1:5 after inactivation of the m copies of SUC genes in the Saccharomyces cerevisiae strain.

### Claim 2 (depends on 1)

The method as claimed in claim 1, wherein the fragment of SEQ ID NO: 3 comprises, or consists of, between 40 and 60 consecutive base pairs of SEQ ID NO: 3.

### Claim 3 (depends on 2)

The method as claimed in claim 2, wherein the fragment of SEQ ID NO: 3 is selected from the group consisting of the sequence consisting of the sequence ranging from base 1 to base 51 of SEQ ID NO: 3, the sequence ranging from base 1549 to base 1599 of SEQ ID NO: 3, the sequence ranging from base 54 to base 104 of SEQ ID NO: 3, the sequence 110 to base 159 of SEQ ID NO: 3, and the sequence ranging from base 1442 to base 1491 of SEQ ID NO: 3.

### Claim 4 (depends on 1)

The method as claimed in claim 1, wherein each of the lox sequences comprises SEQ ID NO: 1.

### Claim 5 (depends on 1)

The method as claimed in claim 1, comprising the following series of steps carried out at least once:
at least two repetitions of the transformation step a) followed by the selection step b); and
a step c) of eliminating the selection markers.

### Claim 6 (depends on 5)

The method as claimed in claim 5, wherein each transformation step a) is carried out with a disruption cassette comprising a different selection marker and/or at least one different homologous recombination sequence.

### Claim 7 (depends on 5)

The method as claimed in claim 5, comprising carrying out said series of steps three times, wherein at least two series comprise four repetitions of the transformation step a) followed by the selection step b).