Plant gene encoding acetyl coenzyme a carboxylase biotin carboxy

Details Plant gene encoding acetyl coenzyme a carboxylase biotin carboxy Plant gene encoding acetyl coenzyme a carboxylase biotin carboxy

1998-01-20

2000-11-21

McElwain, Elizabeth F.

Multicellular living organisms and unmodified parts thereof and

Method of introducing a polynucleotide molecule into or...

The polynucleotide alters fat, fatty oil, ester-type wax, or...

800286, 800298, 435419, 435468, 536 236, A01H 500, C12N 1582, C07H 2104

Patent

active

061505862

DESCRIPTION:

BRIEF SUMMARY
This invention relates to a plant gene specifying the biotin carboxyl carrier protein (BCCP) a subunit of acetyl Coenzyme A carboxylase (ACCase) and to plant genomes genetically transformed with the said gene. Particularly, but not exclusively, the invention relates to ACCase BCCP genes from plants of the Brassica species, especially Brassica napus (oilseed rape) and control of expression of the gene in Brassica plants which are genetically transformed with the gene, a partial gene or its antisense configuration.
Two principal methods for the control of expression are known. These are referred to in the art as "antisense downregulation" and "sense downregulation" or "cosuppression". Both of these methods lead to an inhibition of expression of the target gene. Overexpression is achieved by insertion of one or more than one extra copies of the selected gene. Other lesser used methods involve modification of the genetic control elements, the promoter and control sequences, to achieve greater or lesser expression of an inserted gene.
In antisense downregulation, a DNA which is complementary to all or part of the target gene is inserted into the genome in reverse orientation and without its translation initiation signal. The simplest theory is that such an antisense gene, which is transcribable but not translatable, produces mRNA which is complementary in sequence to mRNA product transcribed from the endogenous gene: that antisense mRNA then binds with the naturally produced "sense" mRNA to form a duplex which inhibits translation of the natural mRNA to protein. It is not necessary that the inserted antisense gene be equal in length to the endogenous gene sequence: a fragment is sufficient. The size of the fragment does not appear to be particularly important. Fragments as small as 40 or so nucleotides have been reported to be effective. Generally somewhere in the region of 50 nucleotides is accepted as sufficient to obtain the inhibitory effect. However, it has to be said that fewer nucleotides may very well work: a greater number, up to the equivalent of full length, will certainly work. It is usual simply to use a fragment length for which there is a convenient restriction enzyme cleavage site somewhere downstream of fifty nucleotides. The fact that only a fragment of the gene is required means that not all of the gene need be sequenced. It also means that commonly a cDNA will suffice, obviating the need to isolate the full genomic sequence.
The antisense fragment does not have to be precisely the same as the endogenous complementary strand of the target gene. There simply has to be sufficient sequence similarity to achieve inhibition of the target gene. This is an important feature of antisense technology as it permits the use of a sequence which has been derived from one plant species to be effective in another and obviates the need to construct antisense vectors for each individual species of interest. Although sequences isolated from one species may be effective in another, it is not infrequent to find exceptions where the degree of sequence similarity between one species and the other is insufficient for the effect to be obtained. In such cases, it may be necessary to isolate the species-specific homologue.
Antisense downregulation technology is well-established in the art. It is the subject of several textbooks and many hundreds of journal publications. The principal patent reference is European Patent No. 240,208 in the name of Calgene Inc. There is no reason to doubt the operability of antisense technology. It is well-established, used routinely in laboratories around the world and products in which it is used are on the market.
Both overexpression and downregulation are achieved by "sense" technology. If a full length copy of the target gene is inserted into the genome then a range of phenotypes is obtained, some overexpressing the target gene, some underexpressing. A population of plants produces by this method may then be screened and individual phenotypes isolated. As with antisense, the inserted

REFERENCES:
patent: 5767362 (1998-06-01), Best

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