The objective of this study was to develop and validate a fast method for typing the main mutations of bovine milk protein genes by using microarray technology. An approach based on the ligation detection reaction (LDR) and a universal array (UA) was used. Polymorphisms in both the coding and noncoding sequences of alpha(S1)-casein, beta-casein, kappa-casein, and beta-lactoglobulin genes were considered because of their well-known effects on milk composition and cheese production. A total of 22 polymorphic sites, corresponding to 21 different variants, were included in the diagnostic microarray. First, a multiplex PCR was developed to amplify all the DNA target sequences simultaneously. Second, the LDR-UA assay was implemented. The method was validated by analyzing 100 Italian Friesian DNA samples, which were also genotyped by conventional methods both at the protein level by means of milk isoelectrofocusing and at the molecular level using PCR-RFLP and PCRsingle strand conformation polymorphism techniques. The genotypes obtained using the LDR-UA approach were in full agreement with those obtained by the conventional analyses. An important result of the LDR-UA assay was a more accurate genotyping of the different milk protein alleles than was found with conventional typing methods. At the kappa-casein gene, in fact, 4 samples were heterozygous (3 reference samples and 1 validation sample) for an allele coding for Thr(136) and Ala(148). This variant, which can be considered as the wild type of the genus Bos, is not usually identifiable by the conventional typing methods used. The multiplex PCR-LDR-UA approach developed provides for an accurate, inexpensive, and high-throughput assay that does not exhibit false positive or false negative signals, thus making it highly suitable for animal genotyping.
机构:
Xuzhou Normal Univ, Sch Life Sci, Key Lab Biotechnol Med Plants Jiangsu Prov, Xuzhou 221116, Peoples R ChinaXuzhou Normal Univ, Sch Life Sci, Key Lab Biotechnol Med Plants Jiangsu Prov, Xuzhou 221116, Peoples R China
Shan, Qun
Zheng, Yuanlin
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Xuzhou Normal Univ, Sch Life Sci, Key Lab Biotechnol Med Plants Jiangsu Prov, Xuzhou 221116, Peoples R ChinaXuzhou Normal Univ, Sch Life Sci, Key Lab Biotechnol Med Plants Jiangsu Prov, Xuzhou 221116, Peoples R China
Zheng, Yuanlin
Chen, Guoqing
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Xuzhou Normal Univ, Sch Life Sci, Key Lab Biotechnol Med Plants Jiangsu Prov, Xuzhou 221116, Peoples R ChinaXuzhou Normal Univ, Sch Life Sci, Key Lab Biotechnol Med Plants Jiangsu Prov, Xuzhou 221116, Peoples R China
Chen, Guoqing
Zheng, Guihong
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Xuzhou Normal Univ, Sch Life Sci, Key Lab Biotechnol Med Plants Jiangsu Prov, Xuzhou 221116, Peoples R ChinaXuzhou Normal Univ, Sch Life Sci, Key Lab Biotechnol Med Plants Jiangsu Prov, Xuzhou 221116, Peoples R China
Zheng, Guihong
Lu, Jun
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Xuzhou Normal Univ, Sch Life Sci, Key Lab Biotechnol Med Plants Jiangsu Prov, Xuzhou 221116, Peoples R ChinaXuzhou Normal Univ, Sch Life Sci, Key Lab Biotechnol Med Plants Jiangsu Prov, Xuzhou 221116, Peoples R China
Lu, Jun
Lv, Xiaojuan
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Xuzhou Normal Univ, Sch Life Sci, Key Lab Biotechnol Med Plants Jiangsu Prov, Xuzhou 221116, Peoples R ChinaXuzhou Normal Univ, Sch Life Sci, Key Lab Biotechnol Med Plants Jiangsu Prov, Xuzhou 221116, Peoples R China
机构:
Univ Calif Los Angeles, Cedars Sinai Med Ctr, Sch Med, Los Angeles, CA 90048 USAUniv Calif Los Angeles, Cedars Sinai Med Ctr, Sch Med, Los Angeles, CA 90048 USA
Kawamata, Norihiko
Ogawa, Seishi
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Univ Tokyo, Sch Med, Tokyo 113, JapanUniv Calif Los Angeles, Cedars Sinai Med Ctr, Sch Med, Los Angeles, CA 90048 USA
Ogawa, Seishi
Yamamoto, Go
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Univ Tokyo, Sch Med, Tokyo 113, JapanUniv Calif Los Angeles, Cedars Sinai Med Ctr, Sch Med, Los Angeles, CA 90048 USA
Yamamoto, Go
Lehmann, Soren
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Univ Calif Los Angeles, Cedars Sinai Med Ctr, Sch Med, Los Angeles, CA 90048 USAUniv Calif Los Angeles, Cedars Sinai Med Ctr, Sch Med, Los Angeles, CA 90048 USA
Lehmann, Soren
Levine, Ross L.
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Harvard Univ, Brigham & Womens Hosp, Sch Med, Boston, MA 02115 USAUniv Calif Los Angeles, Cedars Sinai Med Ctr, Sch Med, Los Angeles, CA 90048 USA
Levine, Ross L.
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Pikman, Yana
Nannya, Yasuhito
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Univ Tokyo, Sch Med, Tokyo 113, JapanUniv Calif Los Angeles, Cedars Sinai Med Ctr, Sch Med, Los Angeles, CA 90048 USA
Nannya, Yasuhito
Sanada, Masashi
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Univ Tokyo, Sch Med, Tokyo 113, JapanUniv Calif Los Angeles, Cedars Sinai Med Ctr, Sch Med, Los Angeles, CA 90048 USA
Sanada, Masashi
Miller, Carl W.
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Univ Calif Los Angeles, Cedars Sinai Med Ctr, Sch Med, Los Angeles, CA 90048 USAUniv Calif Los Angeles, Cedars Sinai Med Ctr, Sch Med, Los Angeles, CA 90048 USA
Miller, Carl W.
Gilliland, D. Gary
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Harvard Univ, Brigham & Womens Hosp, Sch Med, Boston, MA 02115 USAUniv Calif Los Angeles, Cedars Sinai Med Ctr, Sch Med, Los Angeles, CA 90048 USA
Gilliland, D. Gary
Koeffler, H. Phillip
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Univ Calif Los Angeles, Cedars Sinai Med Ctr, Sch Med, Los Angeles, CA 90048 USAUniv Calif Los Angeles, Cedars Sinai Med Ctr, Sch Med, Los Angeles, CA 90048 USA
机构:
INRA, UMR1313, Unite Genet Anim & Biol Integrat, Jouy En Josas, France
AgroParisTech, UMR1313, Unite Genet Anim & Biol Integrat, Jouy En Josas, FranceINRA, UMR1313, Unite Genet Anim & Biol Integrat, Jouy En Josas, France
Rocha, D.
Billerey, C.
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INRA, UMR1313, Unite Genet Anim & Biol Integrat, Jouy En Josas, France
AgroParisTech, UMR1313, Unite Genet Anim & Biol Integrat, Jouy En Josas, France
Univ Paris 11, Inst Genet & Microbiol, UMR8621, Orsay, France
CNRS, Inst Genet & Microbiol, UMR8621, F-91405 Orsay, FranceINRA, UMR1313, Unite Genet Anim & Biol Integrat, Jouy En Josas, France
Billerey, C.
Samson, F.
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机构:
INRA, UR1077, Jouy En Josas, FranceINRA, UMR1313, Unite Genet Anim & Biol Integrat, Jouy En Josas, France
Samson, F.
Boichard, D.
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INRA, UMR1313, Unite Genet Anim & Biol Integrat, Jouy En Josas, France
AgroParisTech, UMR1313, Unite Genet Anim & Biol Integrat, Jouy En Josas, FranceINRA, UMR1313, Unite Genet Anim & Biol Integrat, Jouy En Josas, France
Boichard, D.
Boussaha, M.
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INRA, UMR1313, Unite Genet Anim & Biol Integrat, Jouy En Josas, France
AgroParisTech, UMR1313, Unite Genet Anim & Biol Integrat, Jouy En Josas, FranceINRA, UMR1313, Unite Genet Anim & Biol Integrat, Jouy En Josas, France