真鲷精子诱导漠斑牙鲆减数雌核发育

摘要

Female southern flounder (Paralichthys lethostigma) grow faster than males. Therefore, farming all-female progenies could maximize profits from cultured fish. In the present study, diploid gynogenesis was induced by activating flounder egg development with UV-irradiated (72 mJ/cm2) Pagrosomus major sperm for 3 min in sea water, and then subjecting the eggs to a cold-shock (0-2℃) treatment for 45 min. Cold shock was used to prevent extrusion of the second polar body. In the control treatments, one or more of these steps was omitted to separately assess the effectiveness of UV irradiation and cold shock. When the initial treatment was applied 3 min after hatching (treatment duration, 45 min), the fertilization rate of eggs activated by UV-irradiated heterologous sperm was (37.2 ± 5.1)%, the malformation rate of newly hatched larvae was (8.3 ± 2.5)%, and the hatching rate was up to (9.4 ± 0.71)%. Flow cytometry and chromosome karyotyping were used to examine the ploidy level of larvae experienced to the cold-shock treatment. All of the gynogenetic larvae were diploid (2n = 48), and no haploid or aneuploid phenomena were detected. The duration of hatching of haploid embryos and gynogenetic embryos was significantly different from that of control normal embryos. There were no significant differences in the morphological index between gynogenetic diploids and normal larvae (P>0.05). All of the haploid larvae (100%) showed abnormal external morphology (P<0.05) while normal diploid larvae and gynogenetic diploid larvae showed normal morphologies. In summary, these methods resulted in successful diploid gynogenesis in southern flounder using heterologous sperm. This technique could serve as a useful tool to produce monosex progenies of southern flounder.%采用紫外线(UV)灭活的冷冻真鲷(Pagrosomus major)精子激发漠斑牙鲆(Paralichthys lethostigma)卵子发育.冷冻真鲷精子适宜的UV灭活剂量为72 mJ/cm2,随着处理时间延长,胚胎的孵化率呈现哈特维希效应,其中40 s灭活组胚胎孵化率达到最高[(29士1.9)％],单倍体率100％.利用冷休克法抑制卵子第二极体排出,成功获得雌核发育二倍体仔鱼.冷休克处理温度为0～2℃,处理起始时间3 min、持续时间45 min时,雌核发育二倍体胚胎的孵化率最高,达(9.4士0.71)％,获得批量雌核发育仔鱼.雌核发育仔鱼经流式细胞仪和染色体倍性鉴定,均为二倍体(2n=48),未发现单倍体和非整倍体现象.雌核发育二倍体组和单倍体组与正常二倍体组组孵化时间差异显著,分别历时64 h 59 min和55 h 49 min.雌核发育二倍体仔鱼与正常二倍体仔鱼形态特征差异不显著(P＞0.05),单倍体仔鱼明显畸形.研究表明,采用适宜的紫外线剂量灭活的冷冻真鲷精子可成功诱导漠斑牙鲆雌核发育,获得雌核发育二倍体后代,研究结果可为漠斑牙鲆全雌化苗种生产提供技术和理论依据.