Molecular control of autonomous embryo and endosperm development

Precocious seed development is usually prevented by a series of mechanisms that ensure seed production results from double fertilization. These events are circumvented in natural apomictic plant species that reproduce clonally through seed. Recent advances in molecular genetics using mutagenic approaches in model sexual plant species, such as Arabidopsis and Zea mays, have revealed some of the mechanisms that prevent such precocious seed development. An understanding of these mechanisms may lead to the development of techniques that will allow future crop plant species exhibiting hybrid vigor to be engineered such that their complex genomes can be fixed indefinitely, thereby maintaining high yields. Our current understanding of the mechanisms underlying the processes of reproductive development is discussed in this review.     

Construction and long term preservation of clonal transgenic silkworms using a parthenogenetic strain

For the functional analysis of insect genes as well as for the production of recombinant proteins for biomedical use, clonal transgenic silkworms are very useful. We examined if they could be produced in the parthenogenetic strain that had been maintained for more than 40 years as a female line in which embryogenesis is induced with nearly 100% efficiency by a heat shock treatment of unfertilized eggs. All individuals have identical female genotype. Silkworm transgenesis requires injection of the DNA constructs into the non-diapausing eggs at the preblastodermal stage of embryogenesis. Since our parthenogenetic silkworms produce diapausing eggs, diapause programing was eliminated by incubating ovaries of the parthenogenetic strain in standard male larvae. Chorionated eggs were dissected from the implants, activated by the heat shock treatment and injected with the transgene construct. Several transgenic individuals occurred in the daughter generation. Southern blotting analysis of two randomly chosen transgenic lines VTG1 and VTG14 revealed multiple transgene insertions. Insertions found in the parental females were transferred to the next generation without any changes in their sites and copy numbers, suggesting that transgenic silkworms can be maintained as clonal strains with homozygous transgenes. Cryopreservation was developed for the storage of precious genotypes. As shown for the VTG1 and VTG14 lines, larval ovaries can be stored in DMSO at the temperature of liquid nitrogen, transferred to Grace’s medium during defrosting, and then implanted into larvae of either sex of the standard silkworm strains C146 and w1-pnd. Chorionated eggs, which developed in the implants, were dissected and activated by the heat shock to obtain females (nearly 100% efficiency) or by a cold shock to induce development to both sexes in 4% of the eggs. It was then possible to establish bisexual lines homozygous for the transgene.     

Clonal diversity in high arctic ostracodes

It remains unclear why the majority of parthenogenetic lineages persist for only brief periods of evolutionary time. However, by characterizing their patterns of genetic variation, it is possible to gain insights regarding their evolutionary origin and potential. We examined clonal diversity patterns in high arctic populations of freshwater ostracodes with the goal of clarifying the factors promoting genotypic diversity. Allozyme electrophoresis showed that the three dominant ostracode species in high arctic ponds reproduce via apomictic parthenogenesis that two of these species (Prionocypris glacialis, Candona rectangulata) were both highly clonally diverse, and had allozyme phenotypes suggestive of polyploidy. Scanning microdensitometry confirmed that many clones of P. glacialis at Igloolik were polyploid. In contrast to most other polyploids, clones of P. glacialis seem to be autopolyploids. Although clonal variation in P. glacialis may reflect multiple transitions to parthenogenesis in an undetected sexual population, it seems likely that genomic recombination associated with polyploidy has also played a role in generating local diversity following the transition to parthenogenesis.     

Structural, metabolic and developmental evaluation of ovulated rabbit oocytes before and after cryopreservation by vitrification and slow freezing

The cryopreservation of oocytes is valuable for the preservation of women's fertility and might also be an interesting tool to preserve animal genetic biodiversity but it is not often used because of the very poor fertility recovered after thawing, especially in rabbit species. The objective of our study was to evaluate the effect of slow-freezing and vitrification on the structural integrity of ovulated rabbit oocytes, their ATP contents, and their developmental competence. Results show that, whatever the method is used, cryopreservation has a dramatic effect on the metabolic integrity, the structural integrity, and the developmental ability of the oocytes. Vitrification and slow freezing both impair the rabbit oocytes viability after thawing but the processes act differently. Further studies are needed to improve the cryopreservation techniques in rabbit species. Moreover, we underlined that morphology and maintenance of the structural integrity of the oocytes are not suitable enough to assess the potential for further development of cryopreserved M_II oocytes. The assessment of ATP metabolism allows efficient evaluation of the viability of the frozen or vitrified oocytes. It should be used in addition to parthenogenesis to better assess the potential for further development.     

哺乳动物同性生殖研究历程及应用前景

繁殖是所有生命形式的基础，同性生殖作为一种特殊的繁殖方式存在于自然界中。同性生殖包含孤雌生殖和孤雄生殖，在哺乳动物中尚未被发现。印记基因等表观遗传机制的存在阻碍了同性生殖胚胎发育。目前利用基因编辑已成功培育出双母系小鼠（Mus musculus domesticus）、单母小鼠和双父系小鼠，双母系小鼠和单母小鼠可正常发育并繁殖后代，双父系小鼠可存活48h。本文综述了哺乳动物同性生殖相关技术以及探究历程，总结了同性生殖技术的应用前景与发展方向。     

Female-biased sex allocation in wild populations of the eriosomatine aphid Prociphilus oriens: local mate competition or transgenerational effects of maternal investment?

Several aphid species exhibit female-biased sex allocation. Local mate competition (LMC) has been postulated to be the evolutionary factor of the female-biased sex allocation. We estimated individual sex allocation in the eriosomatine aphid Prociphilus oriens and explained the observed pattern of sex allocation based on a hypothesis other than LMC. On the basis of the relationship between maternal body size and brood size, we estimated the cost of producing a female to be 1.85 times the cost of producing a male. The population-wide allocation to males was 22–24 %. Winged mothers exhibited a large variation in the number of male and female embryos they had, including 23–30 % of winged mothers producing only female embryos. There was polymorphism in the sex-ratio expression. Thus, the constant male hypothesis assuming LMC was not supported. Winged mothers that produced an all-female brood contained larger female embryos than did mothers that produced a bisexual brood. Previous studies have indicated that a large sexual female produces a single large egg, which hatches into a first-instar larva containing a larger amount of gonads. Thus, in eriosomatine aphids, maternal investment in daughters directly affects the potential fecundity of granddaughters, whereas investment in sons does not. We propose a hypothesis that higher fitness returns from maternal investment in daughters than in sons may have primarily led to the evolution of highly female-biased sex allocation in P. oriens.     

The incidence of aneuploidy after single pulse electroactivation of mouse oocytes

A brief electric pulse often produces a high rate of activation of recently ovulated oocytes. Some other efficient parthenogenetic stimuli, such as alcohol, however, disrupt the spindle apparatus and increase the incidence of aneuploidy. In this paper, we have determined whether electroactivation per se increases the incidence of chromosomal segregation errors in haploid parthenogenones as evidenced at first cleavage mitosis. Superovulated F1 hybrid female mice were killed at 15.5, 18.5, 22.5, and 25 h after the HCG injection. Batches of 10–12 cumulus-denuded oocytes were transferred to an electroactivation chamber containing mannitol which was connected to a high voltage pulse stimulator and the pulse was triggered once. A high proportion of oocytes activated following this treatment, but only the single-pronuclear haploid parthenogenones were incubated overnight in medium containing colcemid, to determine the incidence of aneuploidy as evidenced at first cleavage mitosis. “Sham” electroactivation groups were also examined for evidence of activation and aneuploidy as described above. In these cases, cumulus-denuded oocytes were put through the electroactivation chamber but the pulse was not triggered. A further group of oocytes was studied to determine the effect of handling and exposure to hyaluronidase on activation frequency and parthenogenetic pathways. Finally, the spontaneous rate of aneuploidy was examined in fertilised embryos of F1 hybrid female mice × Rb(1.3)1Bnr male mice at first cleavage mitosis. The results show that single pulse electroactivation does not increase the level of aneuploidy in single-pronuclear parthenogenones compared to the “sham” group or the spontaneous rate observed in 1-cell fertilised embryos, nor does aneuploidy appear to increase with postovulatory age. The developmental pathways observed in the electroactivation group are significantly different to those observed in the “sham” group, and the level of activation observed in both groups is increased through handling of oocytes and their exposure to hyaluronidase. © 1993 Wiley-Liss, Inc.     

Evolution of apomixis as a strategy of colonization in the dioecious species <Emphasis Type="Italic">Lindera glauca</Emphasis> (Lauraceae)

I have investigated the reproductive biology of four dioecious species of Lindera in Japan: Lindera obtusiloba Bl., L. umbellata Thunb., L. erythrocarpa Makino, and L. glauca Bl. The sex ratios in populations of the first three species are close to equality, but in L. glauca only female individuals are found in Japan, although males are known from continental Asia. The persistence of this dioecious species in the absence of males is surprising, and prompts the question: What mechanisms operate to ameliorate problems of colonization in species of Lindera? I carried out bagging experiments in order to test for apomixis (asexual reproduction by seeds) and to establish the importance of pollination and fertilization, and potential pollen vectors. Only L. glauca reproduced by apomixis. Level of fruit set was high, and was not affected by pollination. In the other three species, seed set was entirely dependent on pollen transfer, which could be accomplished by small Coleoptera and Diptera. The evolution of apomixis in L. glauca appears to have been a strategy to overcome sterility and establish a population in the Japanese islands even in the absence of males. However, this change in breeding system has not occurred in congeneric, co-occurring species, which do not seem to have experienced a lack of male plants.     

Production of females by parthenogenesis in the ant,Cerapachys biroi

Summary Groups of virginCerapachys biroi workers produced females (workers) in the laboratories. This is the first report of thelytoky in Cerapachyinae.     

Effect of in vitro and in vivo colchicine treatments on pollen production and fruit set of melon plants obtained by pollination with irradiated pollen

Haploid/doubled haploid (DH) technology can aid plant breeding programs by accelerating production of homozygous lines, provided enough viable DH progeny can be obtained from diverse haploid genotypes. In cases where there is a low frequency of spontaneous doubling, chromosome doubling procedures are required to achieve fecundity. We produced 63 parthenogenetic melon plantlets via pollination with γ-irradiated pollen, cloned them by nodal cuttings, and tested the effects of in vitro and in vivo colchicine treatment on survival, ploidy, pollen production, and fruit recovery. The most effective procedure was in vitro exposure of 3 cm shoot tip explants to 500 mg/l colchicine for 3 h. This treatment gave 83% survival of explants and 26% conversion to diploidy. Fruit recovery rate was 60% among plants with good pollen production. In vivo exposure of the tops of young plants to 5000 mg/l for 2 and 4 h yielded some fruits but also resulted in less survival and more morphological abnormalities. Strategies for recovery of progeny from parthenogenetic melon plants are recommended. To our knowledge, this study represents the first comprehensive study of recovery of fruits and viable seeds from parthenogenetic melon plants.