拟穴青蟹Cactus基因的cDNA克隆、序列及生物学功能

为获取拟穴青蟹Cactus基因cDNA全长、分析基本生物学信息,并初步探索其在病原物刺激下的免疫反应,实验采用RACE技术获得了拟穴青蟹Cactus(SpCactus)基因的cDNA全长序列,其cDNA全长为2035 bp,开放阅读框(ORF)1311 bp,编码436个氨基酸,分子量为46.01 ku。对蛋白理化性质进行预测发现,SpCactus为亲水性蛋白,等电点pI为4.91。经预测,SpCactus与其他物种的IκB蛋白具有相似的功能结构域。同源性比对结果显示,SpCactus与凡纳滨对虾和中国明对虾的Cactus蛋白同源性均高达62%,相似度为73%。系统进化树分析显示,SpCactus与甲壳动物聚为一支,与无脊椎动物聚为一大支。实时荧光定量PCR(RT-PCR)检测发现,SpCactus基因在拟穴青蟹不同组织中均有表达,肌肉中的表达量最高,其次为心脏、眼柄、血液和鳃,肝胰腺中的表达量最低。LPS和金黄色葡萄球菌刺激均能显著诱导SpCactus基因的表达。本实验成功扩增了SpCactus基因全长,并进行了生物信息学分析、理化性质预测,初步探讨了其生物学功能,为进一步研究其在免疫反应中的生物学功能提供参考依据。     

The effects of diet supplemented with Lactobacillus rhamnosus on tissue parameters of rainbow trout,Oncorhynchus mykiss (Walbaum)

This study was carried out to establish the effects of a 6 week treatment with the diet supplemented with L. rhamnosus in concentrations of 10⁷ CFU g^?1 (G1 group) and 10⁸ CFU g^?1 (G2 group) on the condition expressed by condition factors (Fulton's, Clark's and B), intestinal microbiology, haematological, histological and selected antioxidative parameters of rainbow trout. A significantly higher condition factors were found in G1 group indicating that higher concentration of probiotic (10⁸ CFU g^?1) did not result in the better condition. Cholesterol and urea levels were significantly higher in both G1 and G2 groups, albumin in G1 and creatinine in G2 group with respect to control. A significantly higher liver TBARS level was observed in G2 group. The feeding with supplemented probiont apparently changed the resident microbiota. Three weeks after withdrawal of the supplemented feed, the microflora mostly reverted to the control composition, although L. rhamnosus in faecal matter of fish remained inherent. The epithelial structure of the proximal and distal intestine revealed the increased absorptive area in both treated groups, as well as the increase in the mucin‐secreting goblet cells. The L. rhamnosus‐treated groups demonstrated the capacity for the augmentation of the innate host defence.     

王氏唇柱苣苔的离体培养及遗传稳定性

以王氏唇柱苣苔(Chirita wangiana)叶片为外植体进行组培离体快繁研究,并利用流式细胞术对组培苗进行遗传稳定性分析。结果表明:最佳初代诱导培养基为MS添加0.5 mg/L 6-BA+0.1 mg/L NAA;最适继代培养基为MS添加0.5 mg/L 6-BA+0.05 mg/L NAA;最适生根培养基为1/2 MS培养基添加0.5 mg/L IBA和15 g/L蔗糖,所有生根培养基获得的组培苗移栽驯化成活率达到92%以上。组织学切片检测表明,王氏唇柱苣苔叶片为外植体所形成芽体均为器官发生方式形成。流式细胞术检测表明,组培苗倍性没有变化,基因组大小与母本相比仅发生了1.86%的减少;染色体数目为2n=36,跟母本一致,植株形态特征上也无变异。研究结果可为王氏唇柱苣苔的园艺应用快速提供大量遗传稳定的种苗。     

柚木繁育技术研究进展

柚木为马鞭草科柚木属落叶或半落叶高大乔木，具有材用、药用、观赏等多种用途。简要介绍柚木的生物学特性及分布现状，综述柚木种子繁殖、扦插繁殖、组织培养技术及栽培管理的研究进展，展望其发展前景，以期为相关研究工作提供参考。     

Effects of fish meal replacement by low‐gossypol cottonseed meal on growth performance,digestive enzyme activity,intestine histology and inflammatory gene expression of silver sillago (Sillago sihama Forsskál) (1775)

A 56‐day feeding trial was conducted to evaluate the effects on the growth performance, digestive enzyme activity, inflammatory genes expression and intestine histology of silver sillago, Sillago sihama (Forsskål 1775), by replacing fish meal (FM) with low‐gossypol cottonseed meal (LCSM). Five isonitrogenous and isolipidic diets were formulated, including R0 group (control, containing 550.0 g/kg FM), R16 group (88.5 g/kg LCSM and 461.5 g/kg FM), R32 group (177.0 g/kg LCSM and 373.0 g/kg FM), R48 group (265.5 g/kg LCSM and 284.5 g/kg FM) and R64 group (354.0 g/kg LCSM and 196.0 g/kg FM). Fish fed R0 and R16 groups had a significantly higher weight gain rate (WGR) and specific growth rate (SGR) than R48 and R64 groups (p < .05). In contrast to whole‐body crude protein, whole‐body moisture increased with the FM level of substitution (p < .05). With the increased amount of LCSM in the diet, the activity of intestinal amylase (AMS) increased significantly (p < .05), and intestinal trypsin (TRP) decreased (p < .05). Dietary LCSM substitution upregulated the expression of intestinal tumour necrosis factor‐α (TNF‐α), the nuclear factor kappa‐light‐chain‐enhancer of activated B cells (NF‐κB), and interleukin one beta (IL‐1β), but downregulated tight junction proteins ZO‐1(ZO‐1), transforming growth factor beta‐3 (TGF‐β3) and interleukin 10 (IL‐10) expression. Histological analysis revealed progressive morphological damage to the mid‐intestine with higher levels of FM replacement. These results showed that 88.5 g/kg (16%) of FM replaced by LCSM with amino acids (methionine and lysine) supplementation did not significantly reduce growth compared with FM‐based control.     

Effects of dietary sodium butyrate on growth,diet conversion,body chemical compositions and distal intestinal health in yellow drum (Nibea albiflora,Richardson)

The present study was conducted to investigate the effects of sodium butyrate (SB) on the physical barrier function, pro‐inflammatory response and possible underlying mechanisms in the distal intestine (DI) of yellow drum when fed a high‐SBM diet. Three iso‐proteic and iso‐lipidic diets were formulated with fish meal (FM, the control), 45% fish meal protein replaced by SBM (SBM) and 45% fish meal protein replaced by SBM but supplemented with 0.15% SB (SBM + SB). Fish were fed twice daily for 10 weeks. Results indicated that SB supplementation significantly increased the specific growth rate (SGR) and feed efficiency ratio (FER) and methionine content of muscle when compared with those of fish fed the SBM diet (p < .05). The morphologic histology results showed that SB dramatically improved the physical barrier structure, characterized as increases of fold height (FH) and muscular thickness (MT) (p < .05). RT‐qPCR data were accordant with morphologic histology results, in which claudin 3, claudin 4 and occludin were increased while claudin 7 and myosin light chain kinase (MLCK) mRNA expression levels were decreased (p < .05). Sodium butyrate also exerted anti‐inflammatory function, which may be attributed to the suppression of nucleus p65 protein expression. Results suggest SB can be incorporated in high‐SBM diets to ameliorate the negative consequences of alternative dietary ingredients on yellow drum physiology.     

氨氮胁迫对凡纳滨对虾肠道免疫相关指标的影响

为研究急性氨氮胁迫对凡纳滨对虾(Litopenaeus vannamei)肠道免疫功能的影响,将对虾暴露于氨氮浓度为20 mg·L^-1的海水中72 h,测定了不同时间点肠道中抗病原感染指标如酸性磷酸酶(ACP)、碱性磷酸酶(ALP)、溶菌酶(Lys)、酚氧化酶原(proPO)以及抗氧化功能指标如总抗氧化能力(T-AOC)、超氧化物歧化酶(SOD)、热休克蛋白70(HSP70)、热休克蛋白90(HSP90)等变化。结果显示,与对照组相比,氨氮胁迫后:1)ACP和ALP活性均于6 h显著升高(P<0.05),随后于48~72 h显著低于对照组(P<0.05);Lys活性于24 h显著升高至最大值(P<0.05),随后于72 h显著低于对照组(P<0.05)。2)T-AOC和SOD活性均于6 h和12 h显著高于对照组(P<0.05),随后于48 h和72 h显著降低(P<0.05)。3)HSP70基因表达水平于24 h显著升高至最大值,随后虽有降低,但仍显著高于对照组(P<0.05);HSP90和proPO基因表达水平均于12 h显著升高至最大值(P<0.05),随后于72 h降低至对照组水平(P>0.05)。研究表明,急性氨氮胁迫对凡纳滨对虾肠道免疫功能相关指标影响显著,对其肠道免疫防御系统有明显的损伤作用。     

草鱼3个6-磷酸葡萄糖酶催化亚基的基因表达分析及高糖饲料对其表达的影响

为了探讨6-磷酸葡萄糖酶催化亚基(glucose-6-phosphatase catalytic subunit,G6PC)在草鱼(Ctenopharyngodon idellus)糖代谢中的作用,采用同源序列比对的方式,在草鱼基因组中获取了3个g6pc基因的序列,通过序列比对和进化树分析,将其分别命名为g6pca、g6pcb1和g6pcb2,其编码的氨基酸序列与斑马鱼(Danio rerio)、虹鳟(Oncorhynchus mykiss)和人(Homo sapiens)具有较高的同源性,相似度分别为85%～94%、64%～83%和54%～66%。同线性分析表明g6pc在草鱼染色体上的分布与斑马鱼等高度相似,表明草鱼g6pc基因在进化中具有较高的保守性。利用RT-PCR检测3个基因在鳃、脂肪、脑、心脏、肝、肾、前肠、中肠、后肠和肌肉10个组织中的表达,结果显示, g6pca在脑和肝中表达量较高,脂肪组织次之; g6pcb1在肝中表达量最高,中肠次之; g6pcb2在心脏表达量最高,其次为脂肪组织。同时探讨了高糖饲料对草鱼不同g6pc亚型转录水平的影响,以及不同浓度葡萄糖和胰岛素刺激草鱼肝细胞(L8824)后对g6pca转录水平的影响。结果显示,饲喂高糖饲料(7周)后,与对照组相比,草鱼肝脏g6pca mRNA水平显著升高, g6pcb1和g6pcb2 mRNA水平无显著变化。离体情况,与5 mmol/L葡萄糖组相比,15 mmol/L葡萄糖显著增加了L8824的g6pca mRNA水平,且1 mol/L胰岛素可以抑制这种作用; 30 mmol/L葡萄糖对L8824 g6pca mRNA水平无显著性影响。本研究表明,草鱼g6pc发生加倍后存在功能分化,高糖可以诱导g6pca mRNA的表达,而对g6pcb1和g6pcb2的转录水平无影响,其具体功能还需进一步研究。     

低盐胁迫下松江鲈HSPB1、HSPB7和HSPB11基因的表达变化规律

为了探究小分子热休克蛋白基因HSPB1、HSPB7和HSPB11在松江鲈(Trachidermus fasciatus)应对低盐胁迫过程中的调节作用,本研究基于前期转录组数据,获取3个目标基因的序列信息并进行了系统进化分析,利用实时荧光定量PCR技术检测了3个基因在两种低盐胁迫处理下不同时间点(0 h、12 h、24 h和48 h)在鳃、肠、肾和肝组织中的表达水平。系统进化分析结果表明, HSPB1、HSPB7和HSPB11基因分别聚类形成独立分支;在各基因分支中,松江鲈与已报道的鲈形目、鲤形目和鳉形目等鱼种共同聚为硬骨鱼类分支。在两种低盐胁迫处理下, 3个基因在鳃组织中的表达量均在12h显著升高,而在肠、肾和肝组织中的表达量则呈现不同的变化趋势。肠组织中,HSPB7和HSPB11在盐度渐变低盐胁迫(盐度变化速率1.1/h)下表达量均显著升高, HSPB1表达量在48 h显著降低;盐度骤变低盐胁迫(盐度变化速率27/h)下HSPB1和HSPB7表达量在24 h显著升高, HSPB11表达量显著降低。肾组织中,HSPB1、HSPB7和HSPB11表达量均仅在盐度渐变低盐胁迫24h显著升高;盐度骤变低盐胁迫下HSPB1表达量显著降低, HSPB7和HSPB11表达量则显著升高。肝组织中, HSPB7无表达; HSPB1表达量在盐度渐变低盐胁迫下无显著变化,但在盐度骤变低盐胁迫下则显著升高;HSPB11表达量在两种处理下均显著升高。本研究比较分析了HSPB1、HSPB7和HSPB11基因在松江鲈应对不同低盐胁迫时表达变化规律的异同,相关结果为探讨小分子热休克蛋白在鱼类应激调节过程中的作用及洄游性鱼类适应盐度变化的分子调控机制提供了理论依据。