Multicellular living organisms and unmodified parts thereof and – Method of introducing a polynucleotide molecule into or... – The polynucleotide alters carbohydrate production in the plant
Reexamination Certificate
1993-08-13
2001-04-10
McElwain, Elizabeth F. (Department: 1638)
Multicellular living organisms and unmodified parts thereof and
Method of introducing a polynucleotide molecule into or...
The polynucleotide alters carbohydrate production in the plant
C800S298000, C800S317200, C435S419000, C435S417000, C435S468000, C536S023600
Reexamination Certificate
active
06215042
ABSTRACT:
This application is a national stage application of PCT/EP92/00302 filed Feb. 11, 1992.
The present invention relates to plasmids containing DNA-sequences which contain information that, after insertion into a plant genome, cause changes in the carbohydrate concentration and the carbohydrate composition in regenerated plants, as well as plant cells and plants containing sequences from these plasmids.
Because of the continual growth in the world population, there is a continually growing demand for nutrient and raw materials. It is the task of biotechnological research to achieve a change in the content as well as the yield of crops. To do this the metabolism of plants has to be altered.
Of particular interest is the possibility of using plant ingredients as renewable sources of raw material sources e.g. for the chemical industry. Renewable sources are of great importance for two reasons. Firstly, up to now, mineral oil and coal deposits have been the main source of raw materials for the petrochemical industry. Such deposits are finite, however, so that alternative, renewable raw material sources must be developed.
Secondly, the present situation of agriculture in Europe and North America has lead to a surplus of crops grown for their nutritive properties. This causes obvious financial and political problems in agriculture. Alternative products for which there is a higher quantitative demand could be a solution to this problem.
Renewable raw materials can be divided into fats and oils, proteins and carbohydrates, such as mono-, di-, oligo- and polysaccharides. The most important polysaccharides are starch and cellulose. In the EEC, the total starch production in 1987-1988 comprised maize (60%), wheat (19%) and potato (21%).
For an increasing use of plant starch as an industrial raw material the quality of the starch must meet the demands of the processing industry. Important considerations include the amylose to amylopectin ratio, the chain length, the branching grade of the amylopectin as well as the size of the starch granules.
The main biochemical synthetic pathways for the production of starch in higher plants are well known. Starch consists of amylose and amylopectin, in which the amylose consists of a linear &agr;-1,4-glucan and amylopectin consists of &agr;-1,4-glucans, which are connected to each other via &agr;1,6-linkages and thus form a branched polyglucan. The so-called branching enzyme (Q-enzyme) is responsible for the introduction of the &agr;-1,6-linkage. One method for the production of starch which only has a linear &agr;-1,4-glucan structure is therefore by the inhibition of the enzymatic activity of the proteins and/or the inhibition of the biosynthesis of the branching enzyme. New biotechnology processes for the genetic alteration of dicotyledonous and monocotyledonous plants by transfer and stable installation of single isolated genes or groups of genes are known (Gasser and Fraley Science 244 1293-1299). The possibility of specific expression of foreign genes inserted in the plant by gene technology, primarily in potato tubers, is also known (EP 375092 and Rocha-Sosa et al., EMBO J. 8, 23-29 (1989)).
The present invention provides plasmids containing DNA-sequences which contain information that, after insertion into a plant genome, cause changes in the carbohydrate concentration and the carbohydrate composition in regenerated plants.
The invention further provides plant cells containing sequences from these plasmids which can be regenerated to whole plants, as well as plants containing sequences from these plasmids.
The term “plant” means a commercially useful plant, preferably maize, barley, wheat, rice, peas, soya beans, sugar cane, sugar beet, tomato, potato or tobacco.
Carbohydrates which can be altered by the DNA sequences are mono-, di-, oligo- or polysaccharides. Starch is an example of a polysaccharide which can be modified in plants and plant cells.
With the plasmids of the invention, it is possible to modify the amylose to amylopectin ratio of the starch in plant cells and in plants. This is possible through the presence of a branching enzyme, located on the plasmid, which has the following sequence: identified as SEQ ID NO:1
10 20 30 40 50 60
1
TCAGGAGCGGTCTTGGGATATTTCTTCCACCCCAAAATCAAGAGTTAGAAAAGATGAAAG
61
GATGAAGCACAGTTCAGCTATTTCCGCTGTTTTGACCGATGACAATTCGACAATGGCACC
121
CCTAGAGGAAGATGTCAACACTGAAAATATTGGCCTCCTAAATTTGGATCCAACTTTGGA
181
ACCTTATCTAGATCACTTCAGACACAGAATGAAGAGATATGTGGATCAGAAAATGCTCAT
241
TGAAAAATATGAGGGACCCCTTGAGGAATTTGCTCAAGGTTATTTAAAATTTGGATTCAA
301
CAGGGAAGATGGTTGCATAGTCTATCGTGAATGGGCTCCTGCTGCTCAGGAAGCAGAAGT
361
TATTGGCGATTTCAATGGTAGGAACGGTTCTAACCACATGATGGAGAAGGACCAGTTTGG
421
TGTTTGGAGTATTAGAATTCCTGATGTTGACAGTAAGCCAGTCATTCCACACAACTCCAG
481
AGTTAAGTTTCGTTTCAAACATGGTAATGGAGTGTGGGTAGATCGTATCCCTGCTTGGAT
541
AAAGTATGCCACTGCAGACGCCACAAAGTTTGCAGCACCATATGATGGTGTCTACTGGGA
601
CCCACCACCTTCAGAAAGGTACCACTTCAAATACCCTCGCCCTCCCAAACCCCGAGCCCC
661
ACGAATCTACGAAGCACATGTCGGCATGAGCAGCTCTGAGCCACGTGTAAATTCGTATCG
721
TGAGTTTGCAGATGATGTTTTACCTCGGATTAAGGCAAATAACTATAATACTGTCCAGTT
781
GATGGCCATAATGGAACATTCTTACTATGGATCATTTGGATATCATGTTACAAACTTTTT
841
TGCTGTGAGCAATAGATATGGAAACCCGGAGGACCTAAAGTATCTGATAGATAAAGCACA
901
TAGCTTGGGTTTACAGGTTCTGGTGGATGTAGTTCACAGTCATGCAAGCAATAATGTCAC
961
TGATGGCCTCAATGGCTTTGATATTGGCCAAGGTTCTCAAGAATCCTACTTTCATGCTGG
1021
AGAGCGAGGGTACCATAAGTTGTGGGATAGCAGGCTGTTCAACTATGCCAATTGGGAGGT
1081
TCTTCGTTTCCTTCTTTCCAACTTGAGGTGGTGGCTAGAAGAGTATAACTTTGACGGATT
1141
TCGATTTGATGGAATAACTTCTATGCTGTATGTTCATCATGGAATCAATATGGGATTTAC
1201
AGGAAACTATAATGAGTATTTCAGCGAGGCTACAGATGTTGATGCTGTGGTCTATTTAAT
1261
GTTGGCCAATAATCTGATTCACAAGATTTTCCCAGACGCAACTGTTATTGCCGAAGATGT
1321
TTCTGGTATGCCGGGCCTTAGCCGGCCTGTTTCTGAGGGAGGAATTGGTTTTGATTACCG
1381
CCTGGCAATGGCAATCCCAGATAAGTGGATAGATTATTTAAAGAATAAGAATGATGAAGA
1441
TTGGTCCATGAAGGAAGTAACATCGAGTTTGACAAATAGGAGATATACAGAGAAGTGTAT
1501
AGCATATGCGGAGAGCCATGATCAGTCTATTGTCGGTGACAAGACCATTGCATTTCTCCT
1561
AATGAACAAAGAGATGTATTCTGGCATGTCTTGCTTGACAGATGCTTCTCCTGTTGTTGA
1621
TGCAGGAATTGCGCTTGACAAGATGATCCATTTTTTTCACAATGGCCTTGGGAGGAGAGG
1681
GGTACCTCAATTTCATGGGTAACGAGTTTGGCCATCCTGAGTGGATTGACTTCCCTAGTG
1741
AGGGCAATAATTGGAGTTATGACAAATGTAGACGCCAGTGGAACCTCGCAGATAGCGAAC
1801
ACTTGAGATACAAGTTTATGAATGCATTTGATAGAGCTATGAATTCGCTCGATGAAAAGT
1861
TCTCATTCCTCGCATCAGGAAAACAGATAGTAAGCAGCATGGATGATGATAATAAGGTTG
1921
TTGTGTTTGAACGTGGTGACCTGGTATTTGTATTCAACTTCCACCCAAATAACACATACG
1981
AAGGGTATAAAGTTGGATGTGACTTGCCAGGGAAGTACAGAGTTGCACTGGACAGTGATG
2041
CTTGGGAATTTGGTGGCCATGGAAGAGCTGGTCATGATGTTGACCATTTCACATCACCAG
2101
AAGGAATACCTGGAGTTCCAGAAACAAATTTCAATGGTCGTCCAAATTCCTTCAAAGTGC
2161
TGTCTCCTGCGCGAACATGTGTGGCTTATTACAGAGTTGATGAACGCATGTCATAAACTG
2221
AAGATTACCAGACAGACATTTGTAGTGAGCTACTACCAACAGCCAATATCGAGGAAAGTG
2281
ACGAGAAACTTAAAGATTCATCATCTACAAATATCAGTACATCATCTACAAAAAATGCTT
2341
ATTACAGAGTTGATGAACGCATGTCAGAAGCTGAAGATTACCAGACAGACATTTGTAGTG
2401
AGCTACTACTACCAACAGCCAATATCGAGGAGAGTGACGAGAAACTTGATGATTCATTAT
2461
CTACAAATATCAGTAACATTGGTCAGACTGTTGTAGTTTCTGTTGAGGAGAGAGACAAGG
2521
AACTTAAAGATTCACCATCTGTAAGCATCATTAGTGATGCTGTTCCAGCTGAATGGGCTG
2581
ATTCGGATGCAAACGTCTGGGGTGAGGACTAGTCAGATGATTGATCGATCCTTCTACGTT
2641
GGTGATCTCGGTCCGTGCATGATGTCTTCAGGGTGGTAGCATTGACTGATTGCATCATAG
2701
TTTTTTTTTTTTTTTTTAAGTATTTCCTCTATGCATATTATTAGCATCCAATAAATTTAC
2761
TGGTTGTTGTACATAGAAAAAGTGCATTTGCATGTATGTGTTTCTCTGAAATTTTCCCCA
2821
GTTTTGGTGCTTTGCCTTTGGAGCCAAGTCTCTATATGTAATAAGAAAACTAAGAACAAT
2881
CACATATATAAAATGTTAGTAGATTACCA.
The property of the branching enzyme to modify the amylose/amylopectin ratio in starch is not limited to a coding sequence exactly as it is shown here but can also be represented by slightly different nucleotide sequences. The property of the branching enzyme is also not changed when the plasmids containing the
Jacobsen Evert
Kossmann Jens
Muller-Rober Bernd
Sonnewald Uwe
Visser Richard G. F.
Frommer Lawrence & HaugLLP
Hoeschst Schering AgrEvo GmbH
McElwain Elizabeth F.
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