الفهرس 
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Abstract
The present work was carried out at Nucleic Acids Research Department (NAR), Genetic Engineering and Biotechnology Research Institute (GEBRI), scientific Research & Technology Applications city, New Borg El-Arab City, Alexandria, Egypt.
The present investigation had been carried out for revealing GH and PRL polymorphism in Holstein cattle reared under Egyptian conditions, looking for association between milk production performance and GH and PRL variants and detection of SNPs of GH and PRL genes and their association with milk production traits in Holstein cattle. The goal of the present study has been achieved by employing different molecular genetic techniques; Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) and DNA sequencing.
Blood samples from Forty eight selected Holstein cattle (24 High and 24 low milk production) were collected into tubes containing an anticoagulant disodium EDTA and then DNA from these samples were extracted.
1- Growth hormone gene polymorphisms
The genomic DNA from 48 animals was used to amplify GH gene which yield a fragment of 329 bp.
Restriction analysis of PCR-RFLP-HapaII of GH gene (329-bp) show three genotypes: HH genotype (224-, 105-bp), GH genotype (329-, 224-, 105-bp) and GG genotype (undigested fragment, 329-bp).
Statistical analysis indicated that genotype GH showed higher milk yield (9893.29 ± 557.69) than GG (8164.93 ± 2070.00) and HH (9311.777 ± 286.89) genotypes and genotype GH had higher least square mean of milk production but without a significant difference between the three genotypes. They may be due to the few number of animals in each genotype.
DNA sequencing of fragment of GH gene (329-bp) revealed nucleotide sequence variation among high and low milk production animals revealed several nucleotide variations. The present experiment showed that animal number 2 with T (SNP1), C (SNP2), G (SNP3), C (SNP4), T (SNP5), A (SNP6), C (SNP7), C (SNP8), G (SNP9) and C (SNP10) nucleotide sequence (115, 195, 226, 244, 249, 251, 261, 264, 267 and 269) for the growth hormone gene can be used as a marker-assisted selection (MAS) to select for high milk production trait. Consequently, these 11 SNP’s markers in bovine GH gene may be useful in genetic improvement of milk production trait in Holstein dairy cattle in general and in particular which reared under Egyptian conditions.
2- Prolactin hormone gene polymorphisms
The genomic DNA from 48 animals was also used to amplify PRL gene which yield a fragment of 294 bp.
Restriction analysis of PCR-RFLP-of PRL gene (249-bp) using different restriction endonucleases: (RsaI - AluI- HaeIII – TaqI- Eco RI – Eco RV – BamHI- HincII- AvaII). Some of these digested the amplified fragment (AluI- HaeIII – TaqI) showing no differences between the different animals under the study. The other restriction enzymes (Rsa1-Eco RI – Eco RV – BamHI- HincII- AvaII) can not digest this fragment in all animals.
DNA sequencing of fragment of PRL gene (294-bp) revealed nucleotide sequence variation among high and low milk production animals. Animal number 3 which consider one of the high milk production animals with G (SNP 1) nucleotide sequence (G226) for the PRL gene, this SNP can be used for marker assisted selection for high milk production to increase and accelerate the rate of genetic improvement on milk production trait.
Many SNP’s has been detected in lower milk producing animals which can be used as a MAS in progeny testing of heifers of lowest milk production.
In conclusion, this work revealed the efficiency of the molecular genetic markers (RFLP and SNPs) in detecting enough GH and PRL polymorphisms in Holstein cattle and to look for association between milk production performance and GH and PRL polymorphisms.
The results were effective in selection for high milk production animals (marker-assisted selection). Also, the analysis of this study confirmed that GH and PRL gene could be a candidate gene for application in marker assisted selection (MAS). The results proved a significant effect of higher least square mean in (GH genotype) of GH gene than other genotypes (HH and GG) but with non significant difference between the three genotypes which may be due to the few number of the animals in each genotype.