الفهرس 
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Abstract
This study was performed at laboratory of Genetics and Genetics engineering, Department of Animal Husbandry and Animal Wealth Development, Faculty of Veterinary Medicine, Alexandria University. Nile tilapia fish were obtained from a private fish farm in El-Riad, Kafr El-sheikh Government.
The aim of this work was to the screen for the genetic variation between large and small body weight Nile tilapia. This has been achieved by DNA sequencing for detection of SNPs at GHR1. Also, through expression analysis of some selected genes related to muscle growth (GHR1, GH, IGF-1, MyoG and MSTN) using qRT- PCR.
Blood samples were collected from 30 monosex male fish (15 large body weight and 15 small body weight) from the same bond at the fishing time into vacutainer tubes containing an anticoagulant (EDTA) and then the DNA were extracted using DNA extraction kits. The GHR1 gene (region 3, 917 bp and region 4, 700 bp) were amplified using polymerase chain reaction technique. These regions were used for screening of the polymorphism of GHR1 gene between large body weight and small body weight fish. The purified PCR products of three large and small body weights were sent for sequencing in one direction using DNA sequencer LGC Company (Germany).
Liver sample which used for RNA isolation, were collected from 9 fish (3 large, 3 medium and 3 small body weight), and were used for screening of the difference in expression of selected genes GHR1, GH, IGF-1, MyoG and MSTN among the large, small and medium body weight fish.
SNPs in GHR1 gene sequence in large and small body weight Nile tilapia was detected compared with accession No. NM_001279601.1. In region 3, fish number 5 has four SNPs at 1663 C>T, 1710 C>G, 2016 C>T and 2200 G>A, and the fish 1, 6 has one SNP at 2161 C>T. In region 4, the fish number 4, 5 and 6 (small body weight) has a repeated SNP at 2200 G>A, this SNP is not detected at the large body weight fish. Fish number 2 (large body weight) has a SNP at 2161 C>T. The SNPs at 2116 C>A, and 2117 A>G appeared in fish number 3 (large body weight) only. A repeated SNP were detected among the three small body weight at 2200 G>A, and 2388 A>T, and can be used as a strong marker in GHR1 gene for culling of small body weight fish. All fish large and small body weight have SNPs at 2501 A>G, 2543 G>A and 2625 G>A.
The presence of SNPs at GHR1 reveal changes in of some amino acid in region 3 and 4. Translation of region 3 of GHR1 gene into 252 amino acids. Amino acid alignment of region 3 among the fish showed change in amino acid number 460 T>S and 562 T>I in fish number 5 (one of small body weight), but other fish of small and high body weight group showed 100 % similarity. The translation of region 4 in all fish revealed presence of 211-213 amino acids. Amino acid sequence alignment of region 4 of GHR1 revealed two amino acid substitutions, 557 R>T in fish 3 (large body weight) and 4 (small body weight), 685L>Q in fish number 2 (large body weight).
Quantitative real time PCR were applied to detect the expression levels of target genes in large, medium, and small body weight. We demonstrate that, the expression of genes related muscle growth (GHR1, GH, IGF-1, MyoG, and MSTN) are variable between the small and large body weight fish
Regarding to body weight there is an increase of GH and MSTN expression in small body weight fish than large body weight fish. On other hand, there is an increase GHR1 and MyoG expression in large body weight fish than small body weight fish. Additionally, there is decreased in expression of IGF-1 in both large and small body weight fish.
As a conclusion, although many SNPs has been detected in GHR1 gene among the large and small body weight Nile tilapia, only two repeated SNPs were detected among the three small body weight (2200 G>A, and 2388 A>T) can be used as a marker assisted selection for culling of small body weight fish.
On the other hand, the SNPs at 2116 C>A and 2117 A>G reported large body weight (Fish number 3). The presence of such SNPs can be used as a marker for selection for increase body weight in Tilapia fish.
The expression profile of myogenic genes (GHR1, GH, IGF-1, MyoG and MSTN) are variable between the small and large body weight fish. The expression of GH and MSTN genes in liver increased in small body weight fish than large body weight fish. GHR1 and MyoG expression increased in in large body weight fish in liver of examined fish indicated to its important role for growth and development trait.