Effect of dietary chitosan on growth performance,haematology, immune response,intestine morphology,intestine microbiota and disease resistance in gibel carp (Carassius auratus gibelio)

A 75‐day experiment was conducted with juvenile gibel carp (Carassius auratus gibelio) (4.80 ± 0.01 g) to evaluate effects of dietary chitosan on fish growth performance, haematology, intestine morphology and immune response. Six isonitrogenous (crude protein: 383 g kg^?1), isolipid (97.5 g kg^?1) and isocaloric (gross energy: 16.7 kJ g^?1) diets were formulated to contain 0, 1800, 4000, 7500, 10 000, 20 000 mg kg^?1 chitosan, respectively. The results showed that the growth was depressed when the fish fed with 10 000 mg kg^?1 chitosan. Serum cholesterol, triglyceride and low‐density lipoprotein decreased in 10 000 and 20 000 mg kg^?1 chitosan. On day 75, blood leucocyte phagocytic activity respiratory burst and alternative pathway of complement haemolytic activity were enhanced in 4000 mg kg^?1 chitosan. The number of goblet cell, intraepithelial lymphocyte of mid‐intestine and microvilli height of distal intestine increased at 4000 mg kg^?1 dietary chitosan. Dietary chitosan modulated intestine microbiota, depressed pathogen bacteria Aeromonas veronii‐like and improved Cellulomonas hominis‐like, Bacillus oceanisediminis‐like and two uncultured bacterium‐like species on day 75. Dietary 7500 and 10 000 mg kg^?1 chitosan enhanced the protection against Aeromonas hydrophila infection. In conclusion, oral administration of dietary 7500 mg kg^?1 chitosan for 75 days is recommended for the survival of gibel carp.     

生物絮团技术对异育银鲫生长性能和抗性的影响

为研究生物絮团技术(Biofloc Technology,BFT)在沿海滩涂鱼类养殖中的应用效果,本实验以滩涂主要养殖种类——异育银鲫(Carassius auratus gibelio)为研究对象,按照BFT养殖模式(BFT组,不换水,只补存蒸发掉或取样部分的水分)和一般养殖模式(对照组,每日换水1次,每次换水1/4~1/3)分别饲养,测定各处理组异育银鲫的生长指标、消化酶活性和免疫相关酶活性,应用实时荧光定量PCR法定量分析热休克蛋白HSP70的相对表达,人工感染试验对比分析BFT养殖模式组和一般养殖模式组异育银鲫生长性能和抗性的变化。结果显示:(1)BFT组异育银鲫增重率、特定生长率和存活率均高于对照组(P0.01),肥满度、脏体比和肝体比与对照组间无显著差异(P0.05);(2)BFT组异育银鲫肠道中淀粉酶、脂肪酶和胃蛋白酶活性显著高于对照组(P0.05),分别提高了53.10%、28.10%和17.99%;(3)BFT组异育银鲫体表黏液中超氧化物歧化酶活性、血清中总抗氧化能力和溶菌酶活性高于对照组(P0.01);(4)BFT组脾、肾、肝和鳃中热休克蛋白HSP70的表达量分别上调了1.29倍、1.34倍、1.87倍和1.68倍;(5)嗜水气单胞菌(Aeromonas hydrophilia)人工感染试验证实,BFT组异育银鲫抗细菌感染能力显著增强。研究表明,BFT养殖模式适于异育银鲫养殖,可促进鱼体生长,增强其应激能力和抗病力。     

蜂胶佐剂对异育银鲫嗜水气单胞菌灭活疫苗的免疫增强试验

本文就蜂胶佐剂对鱼类细菌性疫苗的免疫增强作用进行了首次评价。试验以异育银鲫嗜水气单胞菌灭活疫苗为对象,比较了蜂胶佐剂、铝胶佐剂和无佐剂三种疫苗对银鲫的免疫效果。经抗体监测表明,免疫接种后第2周,蜂胶佐剂组银鲫已检测到血清抗体,随后抗体效价迅速上升,第4周达到峰值2^5：铝胶佐剂组和无佐剂组第3周检测到抗体．铝胶佐剂组第5周达到峰值2^5,无佐剂组第4周达到峰值2^4。银鲫免疫后第5周进行攻毒保护试验,发现蜂胶佐剂组、铝胶佐剂组和无佐剂组的免疫保护率分别为77．3％．65．0％和56．3％。试验结果表明,蜂胶佐剂能有效增强异育银鲫细菌性灭活疫苗的免疫保护力。     

异育银鲫病原温和气单胞菌表型及分子鉴定与溶血素基因检测

2008年10月江苏盐城大丰精养异育银鲫（Carassius auratus gibelio) 发生严重疾病,从病鱼肝脏、血液及腹水中分离到优势生长的细菌。对分离做纯培养的4株菌(JY081016-1至JY081016-4)进行形态特征、理化特性等表型生物学性状检验;测定了菌株（JY081016-1）的16S rRNA和gyrB基因序列,分析了16S rRNA和gyrB两种基因序列的同源性,并构建了系统发生树。结果表明分离菌对供试健康异育银鲫有强致病性,菌株（JY081016-1）所扩增的16S rRNA基因序列长度为1448bp（GenBank 登录号：GQ232759）,所扩增的gyrB基因序列长度为1177bp（GenBank 登录号：GQ232760）,其16S rRNA和gyrB基因序列与GenBank数据库中维氏气单胞菌和维氏气单胞菌温和生物型的16S rRNA和gyrB基因序列相似性均在97%以上;根据分离菌的表型及分子特征,判定分离鉴定的4株菌为温和气单胞菌（Aeromonas sobria）。胞外酶及溶血活性检测表明分离菌均能产生蛋白酶、卵磷脂酶、脂酶,在含7%家兔脱纤血液营养琼脂培养基上呈β型溶血;设计的特异性引物可扩增出溶血素基因。     

The effects of dietary selenium on growth performances,oxidative stress and tissue selenium concentration of gibel carp (Carassius auratus gibelio)

A 100‐day growth trial was executed to determine the dietary selenium (Se) requirement of juvenile gibel carp (Carassius auratus gibelio). Selenomethionine was supplemented to casein‐gelatin diets at 0, 0.1, 0.25, 0.5, 1.0, 2.5 and 5 mg Se kg^?1, respectively. Each of these seven semi‐purified diets containing 0.34, 0.47, 0.66, 0.82, 1.23, 2.77 and 5.13 mg Se kg^?1 was fed to triplicate groups of gibel carp (2.74 ± 0.02 g) in a flow‐through system. No behaviour abnormalities and no mortality were found in fish exposed to dietary Se concentrations. With the increasing dietary Se supplemented concentrations, weight gain of fish remarkably increased at the levels of ≤1 mg Se kg^?1 diet and then showed no significant difference above 1 mg Se kg^?1 levels. Although growth performances (weight gain, hepatosomatic index, condition factor and survival) were not impaired in gibel carp fed at above the levels of 2.5 mg Se kg^?1, indicators of oxidative stress were changed significantly. Serum superoxide dismutase (SOD) activities significantly declined, hepatic glutathione peroxidase (GPx) activities significantly increased and the tissue Se concentrations significantly raised at the highest supplemented level of 5 mg Se kg^?1. A clear linear relationship between Se‐depended GPx activities and hepatic Se concentrations was observed. The present results indicated that the dietary Se requirement for gibel carp is 1.18 mg Se kg^?1 diet based on weight gain, GPx activities and tissue accumulation.     

分级对银鲫生长和个体大小差异的影响

在室内水槽进行12周的饲养实验以检验分级对银鲫生长和鱼群内个体大小差异的影响。用隔网将实验水槽分为两部分,每个水槽内放养10尾大鱼（体重8.0~9.0 g）和10尾小鱼（体重6.5~7.5 g）,分级时将个体大小相近的10尾鱼分布在隔网同一侧。实验中,3组鱼分别隔1周、2周或4周分级1次,1组鱼不进行分级作为对照。鱼摄食率和生长率随分级间隔时间缩短而降低。实验结束时,隔1周分级1次的鱼体重低于对照组,隔4周分级1次的鱼体重与对照组无显著差异。实验结束时被分级的鱼群中大鱼体重的变异系数高于小鱼,表明大鱼对分级操作更敏感。本实验结果表明,分级未能明显提高银鲫生长速度和降低鱼群内个体大小的差异程度。     

Effect of partial replacement of dietary monocalcium phosphate with neutral phytase on growth performance and phosphorus digestibility in gibel carp,Carassius auratus gibelio (Bloch)

A feeding trial was conducted to study the effect of partial replacement of dietary monocalcium phosphate (MCP) with neutral phytase on growth performance and phosphorus digestibility in gibel carp, Carassius auratus gibelio (Bloch). Control diet was prepared with 2% MCP but without phytase (P0). Other three experimental diets were prepared by replacement of MCP by 25%, 50% and 75% respectively in comparison with control with supplementation of neutral phytase at 500 U kg^?1 diet in each and designated as P25, P50 and P75 respectively. Gibel carp (initial body weight of 30.22 ± 1.98 g) were reared in twelve 300‐L cylindrical fibreglass tanks provided with filtered flow‐through tap water at 26–28°C. After 8‐week experiment, gibel carp fed with P50 had no obvious differences from the control group on weight gain (WG), specific growth rate (SGR), feed conversion ratio (FCR), protein efficiency rate (PER) and survival rate. Phytase supplementation did not affect body compositions or muscle compositions. Crude protein and phosphorus (P) contents in the faeces of fish fed with the phytase‐supplemented diets were significantly lower than those of the control group. The apparent digestibility coefficients (ADCs) of crude protein and P in gibel carp were increased when fish fed with the diets in which MCP was replaced by neutral phytase. This study suggested that partial replacement of dietary MCP at 50% with neutral phytase was considered as a recommended dietary supplemental level and increased dietary P and protein availability.     

Response of gibel carp (Carassius auratus gibelio) to increasing levels of dietary lysine in zero fish meal diets

To investigate the response of gibel carp to dietary lysine levels in zero fish meal (FM) diets, 8 experimental diets were designed. Among them, diet 1 and diet 2 with 150 g/kg FM, methionine, lysine and threonine were supplemented in diet 2. Diet 3 was without FM but matched the amino acid profile of diet 2 except for lysine. Diets 4–8 were based on diet 3, with graded levels of lysine. After 8‐week feeding trail, final body weight, weight gain (WG), specific growth rate, feed conversion ratio (FCR) and feed intake were significantly improved by diets 1–2 and 6–8 compared with diet 3. Plasma glucose, triglyceride, immunoglobulin G and aspartate aminotransferase levels in diets 4–8 were higher than those in diet 1. Apparent digestibility coefficients (ADCs) of protein, energy, methionine, lysine, valine, histidine and phenylalanine in diet 1 were significantly lower than those in diet 6. The highest ADC of isoleucine was found in diet 3. TOR, IGF‐1, JAK3, STAT4, STAT6 and PepT2 mRNA levels were significantly increased with increasing dietary lysine up to a certain level in zero FM diet and lower than diets 1–2. Based on WG and FCR, the optimal lysine requirement of juvenile gibel carp was determined to be 24.4 g/kg and 24.2 g/kg (65.9 g/kg and 65.4 g/kg of dietary protein) in zero FM diet, respectively, using quadratic regression analysis.     

Effects of dietary lipid levels on growth performance,whole body composition and fatty acid composition of juvenile gibel carp (Carassius auratus gibelio)

We evaluated the effects of dietary lipid levels on the growth, whole body composition and fatty acid composition of juvenile gibel carp (Carassius auratus gibelio). Triplicate groups of 120 juvenile Carassius auratus gibelio (average weight: 2.05 g) were fed four isonitrogenic diets formulated with lipid levels of 1.4% (low), 6.1% (control), 11.6% (medium) and 21.1% (high) for 60 days. Weight gain in the 11.6% lipid group was significantly higher than that in the other groups (P < 0.05). The feed conversion ratio decreased and protein efficiency ratio increased (P > 0.05) as dietary lipid levels increased. N‐3 polyunsaturated fatty acids (PUFA) were not detected in faeces. The whole body lipid contents of 11.6% and 21.1% lipid level groups were significantly higher than that of the 1.4% and 6.1% lipid level groups (P < 0.05). The content of whole body n‐3 PUFA in the 21.1% lipid level group enhanced significantly (P < 0.05). The whole body contents of eicosapntemacnioc acid (EPA) and docosahexaenoic acid (DHA) in the 21.1% lipid level group were the highest (P < 0.05). These results indicated that high dietary lipid levels (21.1%) inhibit weight gain and promoted fat and n‐3 PUFA deposition in juvenile Carassius auratus gibelio, which led to liver damage. A dietary lipid level of 11.6% was determined to be optimal for growth performance of juvenile Carassius auratus gibelio.     

银鲫crh基因的克隆、组织表达谱及其对摄食的影响

为研究crh (Corticotropin-releasing hormone)基因对鱼类摄食的调控作用,首次克隆了银鲫(Carassius auratus gibelio) crh基因c DNA全长序列。运用实时荧光定量PCR(RT-qPCR)技术检测crh基因m RNA在不同组织的表达及餐前、餐后和禁食对其表达的影响。结果显示,银鲫crh基因的c DNA序列全长为920 bp,其中,5’-UTR为53 bp,3’-UTR为378 bp,开放阅读框(Open reading frame,ORF)为489 bp。推导银鲫crh基因的ORF区编码蛋白由162个氨基酸组成,其中,含有11个氨基酸的保守区,24个氨基酸的信号肽,41个氨基酸的成熟肽。氨基酸序列的多重比较分析显示,银鲫crh基因与金鱼(Carassiusauratus)的同源性为99%,与鲤(Cyprinuscarpio)的同源性为96%。荧光定量PCR表达分析显示,银鲫crh基因在下丘脑中的表达量最高,其次是端脑和心脏。crh基因的表达量在餐前与餐后没有显著变化(P0.05),在禁食第5天、第7天出现极显著降低(P0.01),而在复投喂后第9天、第11天出现极显著升高(P0.01)。以上结果显示,crh基因在银鲫摄食调控方面可能扮演着重要角色。     

Effect of dietary magnesium supplementation on the growth performance of juvenile gibel carp,Carassius auratus gibelio

Gibel carp (Carassius auratus gibelio) of mean initial weight 3.1 g were fed one of seven casein‐dextrin‐based diets containing graded levels of magnesium (Mg) (39, 120, 220, 380, 700, 1600 and 2900 mg kg^?1) for 3 months with the waterborne Mg concentration of 10.6–12.7 mg L^?1. Magnesium sulphate was used as the supplementation Mg source in the diets. The experiment was carried out in a flow‐through system. Growth, survival rate, Na⁺/K⁺‐ATPase, Mg²⁺‐ATPase and tissue mineral contents were measured to investigate the effect of dietary magnesium in gibel carp. At the end of the experiment, the hepatopancreas of fish were collected for enzyme determination. The hepatopancreas, vertebrae and whole body were collected for tissue magnesium content analysis. After 3 months, dietary magnesium supplementation did not improve the growth performance, including feed intake, weight gain and feed conversion efficiency of juvenile gibel carp. On the contrary, negative impacts on survival, reduced growth performance and dramatically decreased Na⁺/K⁺‐ATPase, Mg²⁺‐ATPase and superoxide dismutase activities were observed in gibel carp fed a high Mg diet of 2900 mg kg^?1. Although serum and hepatopancreas Mg and Ca contents were not affected by dietary Mg supplementation, vertebrae and whole‐body Mg contents increased significantly with the increasing dietary Mg concentrations. Based on the relationship between whole‐body Mg retention and dietary Mg concentration, a suitable dietary Mg level of 745 mg kg^?1 could be estimated for gibel carp. It could be concluded that dietary Mg supplementation did not improve the growth performance, but could increase vertebrae Mg contents of gibel carp. Considering the adverse effects, a dietary Mg concentration of above 2900 mg kg^?1 is not recommended and it should be careful to supplement magnesium in practical diets for gibel carp as most feed ingredients contain high magnesium concentrations.     

Effects of dietary vitamin C on growth,gonad development and antioxidant ability of on‐growing gibel carp (Carassius auratus gibelio var. CAS III)

An 88‐day experiment was conducted in a flowing system to evaluate the effects of dietary vitamin C on growth, body composition, antioxidant and gonad development of on‐growing gibel carp. Triplicate tanks of gibel carp (77.2 ± 0.1 g) were randomly fed with one of seven experimental diets containing l ‐ascorbic acid of 0, 101.1, 188.5, 313.1, 444.1, 582.1 and 747.0 mg/kg, respectively. The results showed that specific growth rate (SGR) and feed efficiency (FE) of fish were not affected by dietary l ‐ascorbic acid. Dietary l ‐ascorbic acid of 444.1 mg/kg diet led to low levels (p < .05) of gonadosomatic index (GSI) and hypothalamic gonadotropin‐releasing hormone (GnRH) in on‐growing fish, as well as the early ovarian stages (Developing stages) compared with fish (Maturing stages) from the other groups. Dietary l ‐ascorbic acid supplementation increased (p < .05) the dorsal muscle collagen content, but did not affect the protein or lipid content of dorsal muscle in gibel carp. Plasma total antioxidant capacity (T‐AOC) and superoxide dismutase (SOD) activity increased (p < .05) and then remained unchanged with the increase in dietary l ‐ascorbic acid levels. Dietary l ‐ascorbic acid of 101.1 mg/kg diet improved (p < .05) plasma lysozyme activity of the fish. Broken‐line regression indicated that dietary l ‐ascorbic acid requirement of 77 g gibel carp was 223.3 or 225.0 mg/kg diet based on plasma T‐AOC or SOD activity, which was a little higher than that based on plasma l ‐ascorbic acid concentration (193.2 mg/kg).     

Partial and total replacement of fishmeal with poultry by-product meal in diets for gibel carp, Carassius auratus gibelio Bloch

Triplicate groups of gibel carp Carassius auratus gibelio Bloch (initial body weight: 4.89 g) were fed for 8 weeks at 24.8–30.8°C with nine isonitrogenous and isoenergetic diets. The control diet (F1) used white fishmeal (FM) as the sole protein source. In the other eight diets (F2–F9), 40.5–100% of FM protein was substituted by poultry by‐product meal (PBM) at 8.5% increments. The specific growth rate (SGR), feed efficiency ratio, protein efficiency ratio, protein retention efficiency and energy retention rate for fish fed PBM diets (F2–F9) were all higher, but not always significantly, than those for fish fed F1. All apparent digestibility coefficients for fish fed PBM diets were lower than those for fish fed F1. Fish fed F1 had a significantly higher hepatosomatic index value than fish fed PBM diets (P<0.05). No significant (P>0.05) effect of diet was found in whole‐body moisture and fat content. Whole‐body protein and energy content for fish fed PBM diets were slightly higher than that for fish fed F1. The optimal replacement level of FM by PBM was estimated by second‐order polynomial regression to be 66.5% in protein.     

盐酸多西环素临床治疗异育银鲫嗜水气单胞菌感染的给药方案的制定

通过高效液相色谱法(HPLC法)测定盐酸多西环素在异育银鲫(Carassius auratus gibelio)血清中不同时间点的血药浓度,并结合药物的体外药效学参数,确定该药的临床给药方案。结果显示,盐酸多西环素对嗜水气单胞菌(Aeromonas hydrophila,AH10)的最小抑菌浓度(MIC)为0.4μg/m L,最小杀菌浓度(MBC)为3.2μg/m L,防耐药突变浓度(MPC)为4.8μg/m L,耐药选择窗(MSW)为0.4~4.8μg/m L。盐酸多西环素的抗菌后效应(PAE)约为2 h;在异育银鲫中能够达到良好的抑菌杀菌效果的盐酸多西环素口灌剂量是20 mg/kg,休药期至少为25 d;能够控制耐药菌株产生的盐酸多西环素口灌剂量是80 mg/kg,休药期至少为60 d。     

Effect of replacement of dietary fish meal by meat and bone meal and poultry by-product meal on growth and feed utilization of gibel carp, Carassius auratus gibelio

Triplicate groups of gibel carp Carassius auratus gibelio (initial body weight: 5.25 ± 0.02 g) were fed for 8 weeks at 20–25 °C on five isonitrogenous (crude protein: 400 g kg^?1) and isoenergetic diets (gross energy: 17 kJ g^?1). Meat and bone meal (MBM) or poultry by‐product meal (PBM) were used to replace fish meal at different levels of protein. The control diet contained fish meal as the sole protein source. In the other four diets, 150 or 500 g kg^?1 of fish meal protein was substituted by MBM (MBM₁₅, MBM₅₀) or PBM (PBM₁₅, PBM₅₀). The results showed that feeding rate for the MBM₅₀ group was significantly higher than for other groups except the PBM₅₀ group (P < 0.05). Growth rate in the MBM₁₅ group was significantly higher than that in the control (P < 0.05), while there was no significant difference in growth between the control and other groups (P > 0.05). Feed efficiency and protein efficiency ratio in MBM₅₀ was significantly lower while that in MBM₁₅ was significantly higher (P < 0.05). Replacement of fish meal by MBM at 500 g kg^?1 protein significantly decreased apparent dry matter digestibility (ADC_D) and gross energy (ADC_E) while apparent protein digestibility (ADC_P) was significantly decreased by the replacement of MBM or PBM (P < 0.05). The results suggest that MBM and PBM could replace up to 500 g kg^?1 of fish meal protein in diets for gibel carp without negative effects on growth while 150 g kg^?1 replacement by MBM protein improved feed utilization.     

银鲫肠道内抑制嗜水气单胞菌的潜在益生菌筛选及其特性研究

从养殖的银鲫(Carassius auratus gibelio)肠道中分离纯化得到60株具有不同特征的菌株,以嗜水气单胞菌(Aeromonas hydrophila)为指示菌,点种法初筛得到12株菌株,再通过牛津杯实验筛选出4株对嗜水气单胞菌有明显抑制作用(抑菌圈直径在14 mm以上)的益生菌疑似株,并进行了耐热试验、耐酸试验和人工肠液耐受试验,筛选出最符合益生菌功能特性的菌株YJ-9。该菌株在90℃高温水浴30 min后相对存活率为1.51%,在pH3的液体培养基中4 h内相对存活率为8.37%,活菌数都保持在106 CFU/mL以上,并且可以在人工模拟肠液中缓慢生长。在急性毒力实验中,10倍于实验剂量的菌株YJ-9对银鲫无毒,动物保护实验证明菌株YJ-9能有效增强银鲫对嗜水气单胞菌的免疫抵抗力,相对保护率达到80%。研究结果表明,菌株YJ-9可以作为一个新的饲用微生态制剂候选菌种。     

异育银鲫病原维氏气单胞菌的分离鉴定及药敏试验

从患病的异育银鲫(Carassius auratus gibelio)肝脏中分离得1株优势菌DF-1。将菌DF-1进行人工回感试验,其患病症状同自然发病症状,证明其对鲫有致病性。该菌的形态特征、主要理化特性、16S rRNA序列测定结果及系统发育树的构建均表明其为维氏气单胞菌(Aeromonas veronii)。药敏试验结果表明,该菌对甲哌利福霉素、链霉素、卡那霉素、庆大霉素、四环素、阿洛西林、氟苯尼考等12种药物敏感,对青霉素、苯唑西林、氨苄西林、罗红霉素、克林霉素等15种药物表现出耐药性。     

Effect of biofloc technology on water quality and feed utilization in the cultivation of gibel carp (Carassius auratus gibelio var. CAS III)

This study was designed to use sucrose as carbon source to investigate the effect of biofloc technology on water quality and feed utilization in the cultivation of gibel carp (Carassius auratus gibelio var. CAS III). Three isolipidic and isoenergetic diets were formulated containing graded levels of dietary protein (257.4, 300.7 and 353.4 g/kg). The control group (P34) was fed with 353.4 g/kg protein diet without biofloc. Three biofloc treatments (P24 + B, P29 + B and P34 + B) were fed with the three diets with daily sucrose supplementation and continuous air inflation. The results found that the sucrose supplementation could generate biofloc and increased biofloc volume (BFV, p < 0.05). The concentration of total ammonia nitrogen (TAN), nitrate nitrogen (NO₃‐N) and the total phosphorus (TP) of all tanks decreased in biofloc groups, and the total nitrogen (TN) of P24 + B treatment tanks was significantly lower than that of the control group (p < 0.05). The growth of P24 + B and P29 + B group was similar with that of control group (p > 0.05). Feed conversion ratio (FCR) of P29 + B was lower than P24 + B and P34 + B (p < 0.05). Protein retention efficiency (PRE) of P24 + B group was significantly higher than other treatments (p < 0.05). Phosphorus retention efficiency (PhRE) of P24 + B group was the highest among all treatments. Compared with the control group, P29 + B group had an improved total antioxidant capacity (T‐AOC), superoxide dismutase (SOD) activity and contained the lowest malondialdehyde level (MDA). The present results indicated that low‐protein diet is more suitable for gibel carp in zero‐water exchange biofloc system compared with high‐protein diet.     

方正银鲫微卫星序列的筛选及PCR扩增产物的初步检测

采用小片段克隆法构建了方正银鲫[（Carassius auratus gibelio（Block）]部分基因组文库，并采用非影印的方法将转化子转移到硝酸纤维素滤膜上。用人工合成、并经放射性同位素[γ-^32P]ATP标记的（CA）15探针对构建的基因组文库进行筛选，获得137个阳性克隆，经测序得到微卫星序列130个，其中完美型、非完美型、混合型分别占64．3％、16．9％和18.8％。应用引物设计软件Primer Prmmer5．0进行引物设计，得到引物103对。合成30对引物，随机选取其中5对进行PCR扩增。结果表明，这5对引物均可以稳定扩增，并且观测杂合度均较高。