Establishment Of The Method Of Sex Discrimination And The Female Specific SCAR Marker In Migratory Eels(Anguilla Japonica) Collected At The Yangtze River Estuary

Japanese eel（Anguilla japonica） is a kind of typical case of catadromous fish.It is the most famous and precious species in Asia.It is important to choose brood stock during the artificial reproduction,because it related to whether it is success of reproduction.But it is difficult to determinate sex from appearance.The main purpose of the experiment was to carry on sex identification of migratory eels collected at the Yangtze River Estuary based on the external morphological indicators,serum biochemical markers and SRAP molecular markers.The main research contents were as follows:1、Analysis of morphological index system and discrimination function of male and female migratory eels（Anguilla japonica）collected at the Yangtze River Estuary.17 morphological datas of 60 eel tracked markers were measured.Determination of male and female according to artificial breeding experiment,30 female,8 male and22 Pseudo male eel were found.The principal component analysis of direct measurements separately for 8 male and 30 female and R-cluster analysis of 15indexes was performed after standardization,and the 15 indicators would be carried on single factor analysis of variance.The results showed that:the principal component analysis and R-cluster analysis showed that the differences of individual traits mainly focused on the size of the body and the characteristics of the snout.There are 8 indicators differences between male and female,the body width/body length,body weight/body length^3,body height/body length,pectoral fin length/head length,head length/width of the lips,the vertical distance from the tip of the lips to the posterior margin of the eye/head length,lips length/head length,and eye diameter index（p<0.05）.On this basis,the stepwise discriminant analysis was performed and the discriminant function was established.Y_female=1276.122X₃+366.352X₄+227.44X₇-116.555,Y_male=821.241X₃+235.998X₄+271.728X₇-95.345（X₄=body weight/body length of^3;X₃=body width/body length;X₇=pectoral fin length/head length）.2、Analysis of the differences of serum biochemical indexes and vitellogenin content between male and female migratory eel（Anguilla japonica）collected at the Yangtze River EstuaryThe differences of 14 serum biochemical indexes and vitellogenin content between female and male migratory Anguilla japonica collected at the Yangtze River Estuary were compared,and the function of sex identification on female and male eel was established.The results showed that serum triglyceride（TG）,low density lipoprotein（LDL-C）,and calcium(Ca²⁺)content of the male eels were significantly higher than those of the female eels（P<0.05）,while the albumin（ALB）,and glucose（Glu）of the female eels were significantly higher than those of the male eels（P<0.05）.There were no significant differences between the female and male eels on the contents of Alanine transaminase（ALT）,aspartate aminotransferase（AST）,alkaline phosphatase（ALP）,total bilirubin（T-bil）,total protein（TP）,creatinine（CREA）,total cholesterol（TC）,high density lipoprotein cholesterol（HDL-C）,and inorganic phosphorus（P）were not sex differences（P>0.05）.Vitellogenin content of the female eels was significantly higher than that of the male eel（P<0.05）.Base on the differences of the 14 serum biochemical indexes between female and male eels,two function were established using stepwise discriminant analysis which could be used for sex determination of Anguilla japonica.3、The establishment of SCAR molecular marke and the screening of SRAP marker of migratory eels collected at the Yangtze River EstuaryAccording to male and female migratory eels collected at the Yangtze River Estuary as research material,using SRAP（sequence-related amplified polymorphism）technology and 140 pairs,sex specific expression characteristic bands were gained through screening eels genome.Through the preliminary screening,the F5R2 specific DNA sequence was obtained.After cloning and sequencing,the fragment was 145bp,and the nucleotide homologous sequence of BLAST was not found.According to the sequencing results,the SCAR（sequence characterized amplified regions）specific primers were designed,and the female specific SRAP marker was transformed into a stable and specific SCAR marker.In order to SCAR marker,it was able to identify the sex of eels.In short,the sex identification of the eel can optimize the artificial breeding broodstock selection,and provide the basis for the success of artificial breeding of eels.