Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving nucleic acid
Reexamination Certificate
1999-12-30
2001-06-05
Myers, Carla J. (Department: 1655)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving nucleic acid
C435S091200, C435S810000, C536S023100, C536S024310, C536S024330
Reexamination Certificate
active
06242191
ABSTRACT:
BACKGROUND
Under the current standards established by the United States Department of Agriculture (USDA), beef from bulls, steers, and heifers is classified into eight different quality grades. Beginning with the highest and continuing to the lowest, the eight quality grades are prime, choice, select, standard, commercial, utility, cutter and canner. The characteristics which are used to classify beef include color, texture, firmness, and marbling, a term which is used to describe the relative amount of intramuscular fat of the beef. Well-marbled beef from steers and heifers, i.e., beef that contains substantial amounts of intramuscular fat relative to muscle, tends to be classified as prime or choice; whereas, beef that is not marbled tends to be classified as select. Beef that is classified as prime or choice, typically, is sold at higher prices than beef that is classified into lower quality grades.
Classification of beef into different quality grades is a subjective process which occurs at the packing facility and involves visual inspection of a region between the 12th and 13th rib of a beef carcass by a certified USDA grader. The grader relies on pictures and other objective aids to make his determinations as to color, texture, firmness, and marbling. Unfortunately, the visual appraisal by the grader is costly, labor intensive, and time-consuming. Moreover, the visual appraisal of a beef carcass cannot occur until the animals are harvested.
Currently there are no methods for identifying live cattle that have or that lack the genetic potential to produce beef that is well-marbled. Such information could be used by the cattle producer to channel calves into particular feeding regimens and to meet the requirements of specific marketing programs. Such information could also be used to identify cattle that are good candidates for breeding. Thus, it is desirable to have a method which can be used to assess the beef marbling potential of live cattle, particularly young cattle.
Another characteristic of beef that is desired by consumers is tenderness. Currently there are no procedures for identifying live animals whose beef, if cooked properly, would be tender Currently, there are two procedures which are used by researchers to assess the tenderness of beef after it has been cooked. The first involves a subjective analysis by a panel of trained testers. The second is the Warner-Bratzler shear force procedure which involves an instrumental measurement of the force required to shear steaks, chops, and ground patties of cooked beef. Both methods are costly and time-consuming and, thus, are not practical on a large scale. Consequently, neither procedure is used at a packing facility. Accordingly, it is desirable to have new methods which can be used to identify carcasses and live cattle that have the potential to provide beef that, if cooked properly, will be tender.
SUMMARY OF THE INVENTION
The present invention provides new methods for identifying cattle and beef carcasses that have markers of beef tenderness. The method comprises extracting DNA from a sample obtained from a bovine animal or beef carcass, amplifying the extracted DNA using a primer referred to hereinafter as the “CSTN” primer and low stringency polymerase chain reaction (PCR) conditions to provide a pool of PCR products, and then assaying the pool for the presence or absence of a PCR product of approximately 350 base pairs, a PCR product of approximately 625 base pairs, a PCR product of approximately 675 base pairs, a PCR product of approximately 1450 base pairs, or combinations thereof. Such PCR products are collectively referred to hereinafter as “markers of beef tenderness”. The CSTN primer is a single stranded oligonucleotide of from 8 to 12 nucleotides and having the sequence 5′CGGGCAGG-3′, SEQ ID NO:1. In a preferred embodiment, the CSTN primer has the sequence 5′-CGGGCAGGAT-3′, SEQ ID NO:2. In accordance with the present method it has been determined that beef obtained from cattle or carcasses whose DNA generates at least one of the markers of tenderness is more tender, as assessed by the Warner-Bratzler shear force procedure, than cattle whose DNA does not generate any of these markers of beef tenderness. It has also been determined that beef obtained from cattle whose DNA generates any two of these four markers of tenderness is more tender than beef obtained from cattle whose DNA generates a single marker of tenderness. It has also been determined that cattle whose DNA generates four markers of beef tenderness provide beef that is more tender that cattle whose DNA generates less than four markers of beef tenderness. Thus, the present method is useful for identifying cattle and carcasses whose beef, if cooked properly, will be tender.
The present invention also provides new methods for identifying cattle and carcasses that have markers of beef marbling. The method comprises extracting DNA from a tissue sample from a bovine animal or beef carcass, amplifying the extracted DNA using a primer referred to hereinafter as the “CPN” primer and low stringency PCR conditions to provide a pool of PCR products, and then assaying the pool for the presence or absence of a PCR product of approximately 175 base pairs, referred to hereinafter as the “first marker of beef marbling”, or a PCR product of approximately 600 base pairs, referred to hereinafter as the “second marker of beef marbling”. Preferably, the method comprises assaying for the presence of both the first marker and the second marker of beef marbling. The CPN primer is a single stranded oligonucleotide comprising from 9 to 12 nucleotides and having the sequence 5′GCGCGAACG-3′, SEQ ID NO:3. In a preferred embodiment, the CPN primer has the sequence 5′-GCGCGAACGA-3′, SEQ ID NO:4. In accordance with the present method, it has been determined that beef obtained from cattle whose DNA generates the first marker of marbling has a higher degree of marbling than beef that has been obtained from cattle whose DNA does not generate the first marker of marbling. Cattle whose DNA generates the second marker of marbling in addition to the first marker of marbling provide beef that has an even higher degree of marbling than beef obtained from cattle whose DNA generates only the first marker of marbling.
The present invention also relates to CSTN primers and CPN primers. The present invention also relates to a kit comprising a first container containing a CSTN primer and a second container containing a CPN primer.
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Fluharty Francis
Jackwood Daral J.
Calfee Halter & Griswold LLP
Myers Carla J.
The Ohio State University Research Foundation
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