First successful inducing gynogenesis in Crassostrea angulata by inactivated tetraploid Crassostrea gigas sperm

Artificial induction of gynogenesis is widely practiced in aquatic animals. However, there have been few reports of obtaining adult gynogenetic marine bivalves. This study established a new and effective method for inducing gynogenesis in C. angulata using inactivated tetraploid C. gigas sperm, ultimately obtaining gynogenetic diploids. The optimal induction conditions were as follows: sperm were irradiated with ultraviolet light (254 nm) at an intensity of 1035 mu W/cm(2) for 60 s before fertilization with the eggs. Fifteen minutes post-fertilization, zygotes were treated with cytochalasin B (0.5 mg/L) for a duration of 20 min under a condition of 28 degrees C. Six different ploidy levels (2n, 3n, 4n, 5n, 6n and aneuploid) of larvae were found by karyotype analysis and flow cytometry methods, with a diploid rate of 63.57 +/- 2.80 % and an induction efficiency of 7.47 +/- 0.33 %. At 6 months after fertilization, induced progenies of 4 different ploidy levels (2n, 3n, 4n and aneuploid) were examined from 3 replicate groups. The proportions of diploids, triploids, tetraploids and aneuploids were 31.67 +/- 3.68 %, 57 +/- 3.74 %, 7.67 +/- 0.94 % and 3.67 +/- 0.94 %, respectively. Diploidy was determined by microsatellite analysis, which did not contain any paternal genes, suggesting them as true gynogenetic progenies. The growth traits (shell length, width, height and whole weight) of the gynogenetic diploids did not show any significant difference (P < 0.05) compared to the control diploids. Meanwhile, triploids demonstrated a significant advantage in growth. Gynogenetic diploids underwent sexual differentiation and consisted of females (31.40 +/- 1.52 %), males (67.67 +/- 2.75 %), and hermaphrodites (0.93 +/- 1.31 %). Interestingly, the gonads of diploids developed well and produced functional gametes. This research provides robust technical support for chromosome engineering breeding and the creation of pure lines of bivalves.