Reproduction and breeding最新文献_第4页

SNPs variety of 3-β-hydroxysteroid dehydrogenase 1 (HSD3B1) gene are related to prostate cancer in some Iraqi individuals 3-β-羟基类固醇脱氢酶1 （HSD3B1）基因多态性与伊拉克部分人群前列腺癌相关

Reproduction and breeding

Pub Date : 2025-03-28 DOI: 10.1016/j.repbre.2025.03.004

Zahraa Isam Jameel

There are numerous single nucleotide polymorphisms in the HSD3B1 gene that have been consistently associated with prostate cancer prognosis in clinical studies. HSD3B1 is a crucial gene that regulates androgens. The aim of this study was to identify any association between the HSD3B1 gene and prostate cancer susceptibility or recurrence in Iraqi patients by analyzing the effects of three specific SNPs: rs33937873 (G313A), rs6203 (C338T), and rs1047303 (1245A). The study included 100 healthy controls and 100 Iraqi patients diagnosed with incidental primary prostate cancer. The current study found that the rs33937873, which contains the heterozygote GA of G313A, is an independent risk factor for prostate cancer [odds ratio (OR) = 0.32, 95 % confidence interval (CI): 0.20–0.94, P = 0.00001]. Conversely, the C allele in rs6203 (C338T) is a significant risk factor for the development of PC (OR = 0.48, 95 % CI: 0.31–0.78, P < 0.001). Genetic testing revealed that the rs1047303 CC genotype increases the likelihood of developing PC (OR = 0.48, 95 % CI: 0.26–0.89, P = 0.02). In summary, the current study suggests that these specific polymorphisms hold considerable promise as key genetic markers for predicting which patients are at an increased risk of recurrence or mortality.

HSD3B1基因存在大量单核苷酸多态性，在临床研究中一直与前列腺癌预后相关。HSD3B1是调节雄激素的关键基因。本研究的目的是通过分析三个特异性snp: rs33937873 (G313A), rs6203 （C338T）和rs1047303 （1245A）的影响，确定HSD3B1基因与伊拉克患者前列腺癌易感性或复发之间的任何关联。该研究包括100名健康对照者和100名诊断为偶发原发性前列腺癌的伊拉克患者。本研究发现含有G313A杂合子GA的rs33937873是前列腺癌的独立危险因素[优势比(OR) = 0.32, 95%可信区间（CI）： 0.20-0.94, P = 0.00001]。相反，rs6203中的C等位基因（C338T）是PC发生的重要危险因素(OR = 0.48, 95% CI: 0.31-0.78, P <；0.001)。基因检测显示rs1047303 CC基因型增加PC发生的可能性（OR = 0.48, 95% CI: 0.26-0.89, P = 0.02）。总之，目前的研究表明，这些特定的多态性作为预测哪些患者复发或死亡风险增加的关键遗传标记具有相当大的前景。

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引用次数: 0

Comparison of testicular development of hybrid crucian carp (Japanese white crucian carp ♀ × red crucian carp ♂) in traditional pond, lotus pond and rice paddy 传统池塘、荷塘和稻田杂交鲫鱼（日本白鲫鱼♀×红鲫鱼♂）睾丸发育的比较

Reproduction and breeding

Pub Date : 2025-03-25 DOI: 10.1016/j.repbre.2025.03.002

Weiling Qin, Tao Dai, Jiamin Pi, Yuling Zhou, Yi Zhang, Siyang He, Xiaolong Zhou, Xiangyu Zhao, Qinqin Xiao, Huan Zhong, Yi Zhou

Traditional pond farming is the most important farming mode around the world. Recently, to develop ecological farming, lotus-fish and rice-fish co-culture modes have been widely used in southern China. However, the understanding of these ecological farming modes is insufficient. Preservation of fertility of fishes is crucial for maintenance of germplasm. In the present study, we focus on the testicular development in a widely farmed variety of crucian carp, hybrid crucian carp (Japanese white crucian carp ♀ × red crucian carp ♂, WR) in traditional pond (TP), lotus pond (LP) and rice paddy (RP). The fishes in the three groups had normal morphology of the testis. Analysis of the tissue sections suggested that LP group had higher content of spermatocytes and lower content of spermatids. The analysis of immunofluorescent markers of different cell types in testis indicated that all types of germ cells could be observed in the three groups while higher spermatocytes ratios were found in LP group. Comparison of steroid hormone levels in serum showed that the concentration of testosterone (T) was lowest in the RP group, while the concentration of luteinizing hormone (LH) was higher in the group LP than in the group TP. RNA-seq indicated that genes associated with meiosis and sperm formation tended to up-regulate both in the LP and RP group compared to the TP group. The qPCR results confirmed higher expression of nanog, gpr125, gfra1, dnd, ccna1, vasa, sox9a, and dmrt1 but lower expression of sycp3 in TP. These evidences suggested that the male hybrid crucian carp from the three modes were all fertile with different characteristics of spermatogenesis.

传统的池塘养殖是世界上最重要的养殖方式。近年来，为了发展生态养殖，华南地区广泛采用了莲鱼和稻鱼共养模式。然而，对这些生态养殖模式的认识还不够充分。鱼类育性的保存对种质资源的维持至关重要。本研究主要研究了在传统池塘（TP）、荷塘（LP）和稻田（RP）中广泛养殖的鲫鱼、杂交鲫鱼（日本白鲫鱼♀×红鲫鱼♂，WR）的睾丸发育情况。三组鱼睾丸形态正常。组织切片分析表明，LP组精母细胞含量较高，精母细胞含量较低。睾丸不同细胞类型的免疫荧光标记分析表明，三组均可观察到各种类型的生殖细胞，LP组的精母细胞比例较高。血清类固醇激素水平比较显示，RP组睾酮(T)浓度最低，而黄体生成素（LH）浓度LP组高于TP组。RNA-seq结果显示，与TP组相比，LP组和RP组与减数分裂和精子形成相关的基因均有上调的趋势。qPCR结果证实，TP中nanog、gpr125、gfra1、dnd、ccna1、vasa、sox9a和dmrt1的表达量较高，而sycp3的表达量较低。这些证据表明，三种模式的杂交雄鲫均具有可育性，且具有不同的精子发生特征。

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引用次数: 0

Quantitative proteomic analysis revealed the protein expression characteristics of primary follicle subtypes in zebrafish 定量蛋白质组学分析揭示了斑马鱼初级卵泡亚型的蛋白表达特征

Reproduction and breeding

Pub Date : 2025-03-12 DOI: 10.1016/j.repbre.2025.03.001

Guangjing Zhang , Jing Huang , Liangyue Peng , Jinhui Liu , Wenbin Liu , Wen Fu

Follicles are the basic structure and functional unit of ovaries. In the previous study, we classified primary follicles (PFs) of zebrafish into four subtypes, compared the DNA methylation and RNA expression characteristics among different subtypes follicles, and preliminarily analyzed the molecular mechanism of PFs development. However, the protein expression characteristics of the four subtypes of PFs remain poorly understood. In this study, a total of 9635 proteins were identified in the four subtypes PFs of zebrafish by proteomic analysis. The proteins highly expressed in the PF-ii and PF-iii were enriched in Notch, MAPK, and mTOR signaling pathways. By combined transcriptomic and proteomic analysis screened five key candidate molecules involved in PFs developmental regulation. These results provide reference to the molecular mechanisms underlying follicular development in fish.

卵泡是卵巢的基本结构和功能单位。在之前的研究中，我们将斑马鱼初级卵泡（PFs）分为四个亚型，比较了不同亚型卵泡的DNA甲基化和RNA表达特征，初步分析了PFs发育的分子机制。然而，人们对这四种亚型 PFs 的蛋白表达特征仍然知之甚少。本研究通过蛋白质组分析鉴定了斑马鱼四种亚型PFs中的9635个蛋白质。PF-ii和PF-iii中高表达的蛋白质富含Notch、MAPK和mTOR信号通路。通过联合转录组和蛋白质组分析，筛选出了五个参与 PFs 发育调控的关键候选分子。这些结果为鱼类卵泡发育的分子机制提供了参考。

引用次数: 0

The expression of liver–gonadal axis genes reveals the long–term effects of exogenous estradiol on gonad development and liver function in GIFT tilapia 肝-性腺轴基因的表达揭示了外源雌二醇对GIFT罗非鱼性腺发育和肝功能的长期影响

Reproduction and breeding

Pub Date : 2025-03-08 DOI: 10.1016/j.repbre.2025.02.003

Ye Yuan , Zexun Zhou , Shandong Chen , Yunyun Liu , Yongchun Li , Yan Miao , Shi Wang , Zhongyuan Shen , Lei Zeng , Jun Xiao , Qinbo Qin , Wuhui Li

The aim of this study was to investigate the effects of exogenous estradiol on the expression of the liver–gonadal axis gene in GIFT tilapia. Larval fish (5 dph) were fed a diet with 100 mg kg⁻¹ 17α–ethynylestradiol (EE2) for 30 days and then transferred to a conventional diet. Sex reversal statistics revealed that more than 60 % of the genotypic male fish transformed into pseudofemales. Histological results showed that the oocytes in pseudofemales developed asynchronously and had a prolonged maturation time compared to those in normal female fish. Global expression and DNA methylation analyses revealed that the expression patterns of the gonad and liver genes in pseudofemales were similar to those in XX female fish. Compared to XY male fish, the differentially expressed genes (DEGs) in pseudofemale gonads were enriched in FoxO signaling pathway, oocyte meiosis and other hormone–regulated pathways, which may affect the growth and development of the ovary. Moreover, the DEGs in liver were enriched mainly in protein digestion and absorption, steroid biosynthesis, and some disease and energy metabolism pathways, which may affect liver immunity and energy metabolism. Furthermore, the co-upregulated genes in the liver and gonads were enriched mainly in steroid biosynthesis and glycolysis/gluconeogenesis, whereas the co-downregulated genes were enriched in GnRH secretion and oxytocin signaling pathway. In addition, some genes associated with hormone secretion, growth, metabolism, and immunity in the liver‒gonad axis, such as Oxt, Spoll and Piwi12, were activated, whereas Hox, cyp21a, gas1 and other genes were inhibited. These results indicated that EE2 affects gene expressions in liver and gonad tissues through the hypothalamic–pituitary–gonadal–liver (HPGL) axis.

本研究旨在探讨外源雌二醇对GIFT罗非鱼肝-性腺轴基因表达的影响。将幼鱼（5 dph）投喂添加100 mg kg−1 17α -乙炔雌二醇（EE2）的饲料，饲喂30 d后转投常规饲料。性别反转统计显示，超过60%的基因型雄鱼转化为假雌鱼。组织学结果表明，与正常雌鱼相比，假雌鱼的卵母细胞发育不同步，成熟时间延长。全球表达和DNA甲基化分析显示，假雌鱼的性腺和肝脏基因的表达模式与XX雌鱼相似。与XY型雄鱼相比，假雌生殖腺中的差异表达基因（DEGs）在FoxO信号通路、卵母细胞减数分裂等激素调控通路中富集，可能影响卵巢的生长发育。此外，肝脏中的DEGs主要富集于蛋白质消化吸收、类固醇生物合成以及一些疾病和能量代谢途径，可能影响肝脏免疫和能量代谢。此外，肝脏和性腺的共上调基因主要富集于类固醇生物合成和糖酵解/糖异生，而共下调基因则富集于GnRH分泌和催产素信号通路。此外，肝-性腺轴部分与激素分泌、生长、代谢和免疫相关的基因如Oxt、Spoll、Piwi12被激活，而Hox、cyp21a、gas1等基因被抑制。上述结果表明，EE2通过下丘脑-垂体-性腺-肝脏（HPGL）轴影响肝脏和性腺组织的基因表达。

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引用次数: 0

Exploring spatio-temporal heterogeneity and inter-domain ecological networks of biological community in a marine ranching habitat: Implications for fishery resources conservation 海洋牧场生境生物群落时空异质性及域间生态网络研究：对渔业资源保护的启示

Reproduction and breeding

Pub Date : 2025-02-27 DOI: 10.1016/j.repbre.2025.02.001

Zeng Lei , Chen Guobao , Zheng Tingting , Chen Yuxiang , Yu jie , Li Wuhui

Habitat changes in marine ranching can cause variations in biological resources and community structure. However, the complex inter-domain ecological network (IDEN) in this particular habitat are not well understood. Thus, we employed field surveys and multivariate statistical analyses to explore the spatio-temporal heterogeneity and the IDEN of biological communities within a marine ranching in southern Yintan of Beihai City, China. Our findings indicated that seasonal succession and artificial reefs (ARs) deployment had simultaneous effects on both biotic and abiotic factors, and the former effect was more pronounced. Based on orthogonal partial least squares discriminant analysis (OPLS-DA), various biota (phytoplankton, zooplankton, and nekton) can be broadly categorized into four groups on seasonal and spatial scales. Variable importance projection (VIP) and redundancy analysis (RDA) demonstrated that autumn biomarkers such as Hemidiscus hardmannianus, Oikopleura rufescens, Lagocephalus lunaris were positively correlated with water temperature (WT), salinity (Sal) and inorganic nitrogen (IN), whereas spring biomarkers such as Rhizosolenia robusta, Euconchoecia aculeata, Penaeus penicillatus were positively correlated with dissolved (DO), chemical oxygen demand (COD), and inorganic phosphorus (IP). The IDEN analysis revealed that 23 species such as Chaetoceros densus, Centropages tenuiremis, and Parargyrops edita played a key role in maintaining the stability of the biological communities in the studied sea area. This study can provide novel insights and theoretical supports for the ecological effects analysis and species protection in marine ranching.

海洋牧场生境的变化会引起生物资源和群落结构的变化。然而，这一特殊生境的复杂域间生态网络（IDEN）尚未得到很好的了解。为此，本文采用野外调查和多元统计分析方法，对北海市银坛南部某海洋牧场生物群落的时空异质性和IDEN进行了研究。研究结果表明，季节演替和人工鱼礁布置对生物因子和非生物因子的影响是同步的，且前者的影响更为明显。基于正交偏最小二乘判别分析（OPLS-DA），浮游植物、浮游动物和浮游生物在季节和空间尺度上大致可分为四类。变量重要性投影（VIP）和冗余分析（RDA）结果表明，秋季生物标志物如Hemidiscus hardmannianus、Oikopleura rufescens、Lagocephalus lunaris与水温（WT）、盐度（Sal）和无机氮（IN）呈正相关，而春季生物标志物如rhizzosolenia robusta、Euconchoecia aculeata、Penaeus penicillatus与溶解（DO）、化学需氧量（COD）和无机磷（IP）呈正相关。IDEN分析结果显示，在研究海域，毛囊毛藻（Chaetoceros densus）、细毛毛藻（Centropages tenuiremis）、编辑副毛藻（Parargyrops edita）等23种生物在维持生物群落稳定方面发挥了关键作用。该研究可为海洋牧场生态效应分析和物种保护提供新的见解和理论支持。

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引用次数: 0

Therapeutic potential of carnitine and N-Acetyl-Cysteine supplementation on sperm parameters and pregnancy outcomes in idiopathic male infertility: A systematic review and meta-analysis of randomized control trials 补充肉碱和n -乙酰半胱氨酸对特发性男性不育症精子参数和妊娠结局的治疗潜力：随机对照试验的系统回顾和荟萃分析

Reproduction and breeding

Pub Date : 2025-02-19 DOI: 10.1016/j.repbre.2025.02.002

Yazan Ranneh , Mohammed Hamsho , Abdulmannan Fadel , Hiba Mahgoub Ali Osman , Elshazali Widaa Ali , Nahla Hwaitalla Mohammed Kambal

This systematic review and meta-analysis aimed to evaluate the impact of L-carnitine (LC), L-acetyl-carnitine (LAC), and N-acetyl-cysteine (NAC) on sperm parameters and pregnancy outcomes in men with idiopathic infertility. A comprehensive search was conducted in PubMed, Embase, Scopus, and Google Scholar databases up to January 31, 2024. Of the 2293 initial citations, 109 studies were eligible, and 14 RCTs involving 1453 men were included in the analysis. LC supplementation significantly increased the sperm concentration (MD = 6.85), total motility (MD = 10.41 %), and normal morphology (MD = 1.78 %). LAC supplementation improved total motility (MD = 17.03 %) and forward motility (MD = 13.5 %), whereas LC + LAC supplementation showed no significant effects. Carnitine combinations (CCs) increased sperm count (MD = 16.75) and volume (MD = 0.19 mL), with improved forward motility (MD = 7.70 %). NAC supplementation enhanced sperm concentration (MD = 3.09) and normal morphology (MD = 1.43 %) and volume (MD = 0.61 mL). LC and CC significantly reduced follicle-stimulating hormone (FSH) and luteinizing hormone (LH) levels. Meta-analysis demonstrated significant heterogeneity in many comparisons. LC, CCs, and NAC may improve sperm parameters in idiopathic infertile men, including sperm concentration, motility, morphology, and volume as well as pregnancy outcomes. However, the overall quality of the included studies was low and high heterogeneity was observed. Future well-designed RCTs with longer durations and fixed doses are warranted to confirm these findings and to assess their impact on pregnancy outcomes.

本系统综述和荟萃分析旨在评估l-肉碱（LC）、l-乙酰-肉碱（LAC）和n -乙酰-半胱氨酸（NAC）对男性特发性不育症患者精子参数和妊娠结局的影响。全面检索PubMed、Embase、Scopus和b谷歌Scholar数据库，截止到2024年1月31日。在2293项初始引用中，109项研究符合条件，14项随机对照试验（rct）包括1453名男性纳入分析。添加LC显著提高了精子浓度（MD = 6.85）、总活力（MD = 10.41%）和正常形态（MD = 1.78%）。补充LAC可改善小鼠的总运动性（MD = 17.03%）和正向运动性（MD = 13.5%），而LC + LAC补充无显著影响。左旋肉碱联合用药（CCs）可增加精子数量（MD = 16.75）和体积（MD = 0.19 mL），改善向前运动（MD = 7.70%）。NAC的添加使精子浓度（MD = 3.09）、形态（MD = 1.43%）和体积（MD = 0.61 mL）恢复正常。LC和CC显著降低卵泡刺激素（FSH）和黄体生成素（LH）水平。荟萃分析显示在许多比较中存在显著的异质性。LC、cc和NAC可以改善特发性不育男性的精子参数，包括精子浓度、活力、形态和体积以及妊娠结局。然而，纳入研究的总体质量较低，异质性较高。未来设计良好、持续时间更长、剂量固定的随机对照试验有必要证实这些发现，并评估其对妊娠结局的影响。

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引用次数: 0

Impact of photoperiod and temperature on melatonin, growth hormone, estradiol, and vitellogenin levels in female African catfish (Clarias gariepinus) reproduction cycle 光周期和温度对非洲鲶鱼生殖周期中褪黑素、生长激素、雌二醇和卵黄蛋白原水平的影响

Reproduction and breeding

Pub Date : 2025-01-24 DOI: 10.1016/j.repbre.2025.01.002

Epro Barades , Iskandar , Ibnu Dwi Buwono , Yuli Andriani

This study aimed to investigate the hormone levels of melatonin (Mel), estradiol (E2), vitellogenin (VTG), and growth hormone (GH) in inducing reproduction cycles in female African catfish, Clarias gariepinus, to develop actionable strategies that directly contribute to the efficiency and sustainability of African catfish farming. The treatments involved in this study are three photoperiod variations (L8:D16, L4:D20, and L0:D24) combined with two different temperatures (28 °C and 32 °C) during 90 days of culture (doc). Serum hormone levels were measured using ELISA, and egg diameter was measured using a microscope every 30 days. The results showed that the biological rhythm of the reproductive cycle of African catfish was accelerated by constant exposure to a photoperiod of L0:D24 and 28 °C for 30 days. In this condition, the levels of hormones involved in the reproductive such as Mel (89.82 ± 5.49 ng/mL), E2 (1.66 ± 0.02 ng/mL), VTG (100.96 ± 0.27 ng/mL) and GH (0.33 ± 0.02 ng/mL) with an average egg diameter of 1.15 ± 0.07 mm. These results highlight the complex interplay between photoperiod, temperature, and reproductive physiology in African catfish, suggesting that environmental manipulation could be a valuable tool for optimizing breeding conditions in aquaculture. The conclusion of this study is that manipulating the photoperiod and temperature is an effective and economical approach to stimulate fish spawning. These findings have important implications for African catfish breeding practices, as they provide a clear and actionable strategy for improving reproductive performance.

本研究旨在研究雌性非洲鲶鱼（Clarias gariepinus）中褪黑激素（Mel）、雌二醇（E2）、卵黄原蛋白（VTG）和生长激素（GH）在诱导繁殖周期中的激素水平，以制定可操作的策略，直接促进非洲鲶鱼养殖的效率和可持续性。本研究涉及的处理是三个光周期变化（L8:D16， L4:D20和L0:D24）结合两种不同的温度（28°C和32°C）在90天的培养（doc）。ELISA法测定血清激素水平，显微镜下每30天测定一次卵直径。结果表明，在L0:D24和28°C的光照条件下持续照射30 d，非洲鲶鱼生殖周期的生物节律加快。在此条件下，蛋蛋平均直径为1.15±0.07 mm, Mel（89.82±5.49 ng/mL）、E2（1.66±0.02 ng/mL）、VTG（100.96±0.27 ng/mL）、GH（0.33±0.02 ng/mL）等与生殖有关的激素水平均达到较高水平。这些结果突出了非洲鲶鱼的光周期、温度和生殖生理之间复杂的相互作用，表明环境操纵可能是优化水产养殖养殖条件的一种有价值的工具。本研究的结论是，控制光周期和温度是刺激鱼类产卵的有效和经济的方法。这些发现对非洲鲶鱼的繁殖实践具有重要意义，因为它们为提高繁殖性能提供了明确和可操作的策略。

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引用次数: 0

Transcriptomic profiling reveals the mechanism of fast growth of an allodiploid hybrid fish 转录组学分析揭示了异源二倍体杂交鱼快速生长的机制

Reproduction and breeding

Pub Date : 2025-01-24 DOI: 10.1016/j.repbre.2025.01.001

Ziyi Huang , Siyang Huang , Hongwen Liu , Bei Li , Jianming Yu , Xuanyi Zhang , Fanglei Liu , Lujiao Duan , Qingfeng Liu

Distant hybridization is an important technique and widely used in fish genetic breeding. In previous research, we obtained the allodiploid hybrid (BR, 2n = 49) derived from the inter-subfamily hybridization of female blunt snout bream (Megalobrama amblycephala, BSB, 2n = 48) and male rare gudgeon (Gobiocypris rarus, RG, 2n = 50), which had obvious growth advantages over the male parent RG. However, less is known about the potential mechanisms of fast growth in BR. In this study, the liver transcriptomes of BR and parents were analyzed by RNA-seq technology. A total of 4.73 × 10⁸ clean reads and 11,040 co-expressed unigenes were obtained. The results showed that the differentially expressed genes in BR were asymmetric compared with those in both parents. Interestingly, the genome-wide expression level dominance (ELD) was biased toward the BSB genome in BR, even if BR inherited one more chromosome from RG than that from BSB. The KEGG analysis of ELD genes showed that most metabolism pathways were enriched in ELD-BSB, such as biosynthesis of unsaturated fatty acids, fatty acid elongation, pyruvate metabolism, and so on. Furthermore, the growth genes of BR were most enriched in the ELD-BSB (34.38 %) than in other categories. The expression levels of some key hormone genes (ghra, ghrb, igf1, igfals, igf2bp2, and grb2) were upregulated in BR compared with those in RG. This study provided an insight into the potential molecular mechanism of the fast growth in BR, which will be of great significance in fish genetic breeding.

远缘杂交是鱼类遗传育种中广泛应用的重要技术。在前期研究中，我们获得了雌性钝口鲷（Megalobrama amblycephala， BSB, 2n = 48）与雄性稀有鲟（Gobiocypris rarus， RG, 2n = 50）亚家族间杂交获得的异源二倍体杂种（BR, 2n = 49），该杂种比父本RG具有明显的生长优势。然而，人们对BR快速生长的潜在机制知之甚少。本研究采用RNA-seq技术分析了BR和父母的肝脏转录组。共获得4.73 × 108个clean reads和11040个共表达unigenes。结果表明，BR的差异表达基因与双亲的差异表达基因不对称。有趣的是，即使BR比BSB多继承一条染色体，BR的全基因组表达水平优势（ELD）也偏向于BSB基因组。对ELD基因的KEGG分析表明，ELD- bsb富集了大部分代谢途径，如不饱和脂肪酸的生物合成、脂肪酸延伸、丙酮酸代谢等。此外，BR生长基因在ELD-BSB中富集最多（34.38%）。与RG相比，BR中一些关键激素基因（ghra、ghrb、igf1、igfals、igf2bp2和grb2）的表达水平上调。该研究揭示了BR快速生长的潜在分子机制，对鱼类遗传育种具有重要意义。

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引用次数: 0

Cloning, expression, and localization of Tekt1 in sterile allotriploid crucian carp Tekt1在不育异体三倍体鲫鱼中的克隆、表达和定位

Reproduction and breeding

Pub Date : 2025-01-08 DOI: 10.1016/j.repbre.2024.12.002

Shuxin Zhang , Liran Zhang , Faxian Yu, Xinge Ouyang, Haoxiang Yang, Qining Zuo, Yujie Huang, Xin Chen, Shengnan Li, Min Tao

Tektins (TEKTs) are constitutive proteins of microtubules associated with flagella, cilia, basal bodies, and centrioles. As one of the testis-specific candidate markers, Tekt1, the first identified member of the TEKT family in mammals, is intimately linked to the formation of sperm flagella and may play a pivotal role in flagellar stability and sperm motility. However, studies on Tekt1 in fish species are still relatively understudied. In this study, the full-length cDNAs of Tekt1 were respectively 1727 bp and 1696 bp in allotriploid crucian carp and diploid red crucian carp, which both comprised a 1209 bp open reading frame (ORF) encoding 402 amino acids. Conversely, the diploid common carp possessed a cDNA length of 1771 bp, characterized by a 1218 bp ORF encoding 405 amino acids. The Western blot analysis revealed that the expression level of Tekt1 protein in the testes of sterile allotriploid crucian carp was markedly decreased in comparison to that of fertile diploid red crucian carp during both pre-spermiation and spermiation periods. The immunohistochemistry analysis revealed abnormalities in the spermiogenesis of allotriploid crucian carp, showcasing a distinctive localization pattern of Tekt1 exclusively present in spermatids, in contrast to diploid red crucian carp, where Tekt1 was detected in both spermatids and spermatozoa. Taken together, these findings suggested differential expression of Tekt1 during spermiogenesis between allotriploid and diploid species, and indicated that the decreased expression of Tekt1 protein in allotriploid crucian carp might be associated with male sterility. Furthermore, these results pave the way for further exploration of reproductive characteristics in male allotriploid crucian carp and offer a theoretical foundation for future research on polyploid breeding.

tektin （TEKTs）是与鞭毛、纤毛、基小体和中心粒相关的微管的组成蛋白。Tekt1是最早在哺乳动物中发现的TEKT家族成员，作为睾丸特异性候选标记之一，与精子鞭毛的形成密切相关，可能在鞭毛稳定性和精子运动中起关键作用。然而，对Tekt1在鱼类中的研究仍然相对不足。本研究在异体三倍体鲫鱼和二倍体红鲫鱼中，Tekt1基因的cdna全长分别为1727 bp和1696 bp，组成1209 bp的开放阅读框（ORF），编码402个氨基酸。相反，二倍体鲤鱼的cDNA长度为1771 bp，其中一个1218 bp的ORF编码405个氨基酸。Western blot分析结果显示，与可育二倍体红鲫相比，不育异体三倍体鲫睾丸中Tekt1蛋白的表达水平在精子形成前和精子形成期均显著降低。免疫组织化学分析显示异体三倍体鲫鱼的精子发生异常，显示Tekt1独特的定位模式只存在于精子中，而二倍体红鲫鱼的Tekt1在精子和精子中都被检测到。综上所述，这些发现提示Tekt1蛋白在异体三倍体和二倍体物种精子发生过程中的表达存在差异，并提示Tekt1蛋白在异体三倍体鲫鱼中表达降低可能与雄性不育有关。本研究结果为进一步探索雄性异体三倍体鲫鱼的生殖特性奠定了基础，并为今后多倍体育种研究提供了理论基础。

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引用次数: 0

Highly efficient disruption of tyrb gene using CRISPR/Cas9 in grass carp (Ctenopharyngodon idella) 利用CRISPR/Cas9高效破坏草鱼tyrb基因

Reproduction and breeding

Pub Date : 2025-01-04 DOI: 10.1016/j.repbre.2024.12.003

Pengfei Zhao , Jiaxiang Cheng , Liang Zhang , Wenbo Li, Shengfei Dai, Minghui Li, Deshou Wang, Xingyong Liu

Grass carp (Ctenopharyngodon idella) is the most important economic freshwater fish species in China. The stable production of high-quality grass carp depends significantly on excellent germplasm. In recent years, the generation of new germplasm based on genome editing has been applied to various cultured fish species. However, until now, there has been very few reports on the application of genome editing technology in grass carp. In this study, one-cell-stage embryos of grass carp were acquired through hormone-induced artificial spawning, thereby enabling the performance of genome editing in this species. The tyrb gene was isolated and chosen as the target of CRISPR/Cas9, because of its easily observable phenotype in F0 mutants. RT-PCR results indicated a high expression level of the tyrb gene in both skin and fin tissues. Subsequently, after the microinjection of the guide RNA (gRNA) and Cas9 protein mixture, targeted mutations were successfully identified through Sanger sequencing. Phenotypic analysis of the F0 mutants revealed that the disruption of tyrb led to a distinct golden phenotype, accompanied by a reduction or even absence of melanophores. Moreover, our data demonstrated that the combined utilization of two or three gRNAs caused large DNA fragment loss and a higher mutation rate in the F0 generation. Overall, this represents an application of CRISPR/Cas9 genome-editing technology in grass carp and may hold great significance for the future generation of new golden grass carp germplasm.

草鱼（Ctenopharyngodon idella）是中国最重要的淡水经济鱼类。优质草鱼的稳定生产在很大程度上取决于优良的种质资源。近年来，基于基因组编辑的新种质的产生已应用于各种养殖鱼类。然而，到目前为止，基因组编辑技术在草鱼上的应用报道很少。在本研究中，通过激素诱导人工产卵获得草鱼的单细胞期胚胎，从而能够在该物种中进行基因组编辑。tyrb基因在F0突变体中容易观察到表型，因此被分离出来作为CRISPR/Cas9的靶基因。RT-PCR结果显示，tyrb基因在皮肤和鳍组织中均有高表达。随后，将gRNA和Cas9蛋白混合物显微注射后，通过Sanger测序成功鉴定出靶向突变。对F0突变体的表型分析显示，tyrb的破坏导致了明显的金色表型，伴随着黑素细胞的减少甚至缺失。此外，我们的数据表明，在F0代中，两种或三种grna的联合利用导致了大量的DNA片段丢失和更高的突变率。综上所述，这是CRISPR/Cas9基因组编辑技术在草鱼中的一次应用，对未来培育新的金草鱼种质具有重要意义。

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