We present a novel way of estimating embryo lethals using doubled haploid (dihaploid or DH) female gametophytes grown in tissue culture. This DH-based lethals estimate is hypothesized to be lower than the more common method where lethals are estimated from one generation of selfing. The chief reasoning is the early mortality spike observed for selfed embryos of larch and other genera in the Pinaceae. This early mortality spike occurs at the stage when diploid embryos embed into haploid female gametophyte tissue. However, this stage is missing for DH embryos which grow in a petri dish thus the DH lethals estimate is hypothesized to be lower compared to seed counts. This DH estimate is based on 20,000 haploid female gametophytes cultures from which two surviving diploidized embryos were recovered. Lethals based on diploidized haploids (DH) have not been reported for a conifer previously because, aside from larch, only a few conifer species continue female gametophyte development in absence of pollen. Our chief finding is that, contrary to our hypothesis, the DH-based lethals estimate was similar to lethals estimated obtained from selfed seeds in published studies. Thus doubling haploid female gametophytes can be used to estimate embryo lethals when diploidization is conducted on a massive scale.

An adult tree of Larix decidua Mill., European larch, was produced from doubling one haploid female gametophyte. Whether this tree can produce normal male meiocytes is the crucial question. This adult’s pollen mother cells (PMCs), or male meiocytes, were squashed and stained. Male meiosis was normal and no abortive pollen grains were observed. This female gametophytic apomict of a conifer, a dihaploid adult, is 100% homozygous yet also reproductively competent with normal male meiosis and functional male pollen. Here we show that doubled female gametophytes can produce embryos and reproductively competent adult trees. This shows a way to gain rapid homozygosity and produce completely inbred lines for larch. This is a novel breeding shortcut reported for the first time for a conifer species.