Application of germplasm preservation in breeding programs for molluscan shellfish aquaculture and restoration

Germplasm are the genetic materials of germ cells, including gametes, embryos, or larvae. Preservation of germplasm is usually achieved through cryopreservation. The technology of cryopreservation has been applied for human artificial reproduction as a clinical treatment for infertility and for livestock as a tool for breeding programs worldwide. For fish and shellfish, cryopreservation has been studied in more than 200 species for preservation of natural resources and conservation of endangered species. For molluscan shellfish aquaculture, this technology can have the following potential applications: 1) Preservation of specific lines or strains. Ongoing breeding programs have yielded specific strains and lines, such as disease resistant oysters. Cryopreservation can be used to preserve these valuable strains and provide gametes for assistance of breeding programs. 2) Preservation of natural wild populations. The cryopreserved germplasm of natural populations will act as a repository of genetic diversity and allow for the continued adaptive genetic variation for aquaculture populations through infusion of new material from wild populations. In addition, a germplasm repository of wild populations can provide easy access as study materials for researchers. 3) Creation of self-fertilization inbred lines. Inbred lines are one of the most valuable resources for breeding programs but difficult to produce and require years of repeated crossing of brothers and sisters or backcrossing. Most bivalves are protandrous, beginning life as males and changing into females as they age. Therefore, with the techniques of non-lethal sperm collection and cryopreservation, self-fertilized lines can be created by using cryopreserved sperm and oocytes from the same individual after sex reversal; 4) Preservation of sperm from tetraploid oysters. Triploid-tetraploid technology is probably the most promising in oyster aquaculture because of the superior traits of triploids. Cryopreservation of sperm from tetraploids can extend the commercialization of triploid-tetraploid technology by the sale of frozen sperm and provide cost savings and security for maintaining tetraploids. In addition, cryopreservation of tetraploids produced each year can offer benefit for maintenance of the tetraploid populations. 5) Assistance for creation of mutant lines. Mutant breeding is an effective approach for creation of new strains or lines but seldom used in animal breeding programs because of its low efficiency. Recently, a new approach called TILLING (Targeting Induced Local Lesions in Genomes) was developed in plants for creation of mutant lines. To use this new technique for animal mutant breeding, sperm cryopreservation is an absolutely required technique. So far, this approach has been applied to zebrafish and puffer fish. With the genome sequencing accomplished in oysters, establishment of mutant lines by TILLING and sperm cryopreservation will benefit aquaculture and research on oyster functional genetics, and 6) Assistance of aquaculture hatchery practice for regular and hybrid seed production. The cryopreserved germplasm materials can function as a reservoir to meet the need for regular and hybrid seed production. For example, the hard clam hybrid offspring of Mercenaria mercenaria with Mercenaria campechiensis showed fast growth and higher survival.