Peruvian scallop X bay scallop hybrid three-line breeding system and method

Abstract

The current invent relates to the construction and application of a three-line breeding system in the Peruvian scallop×bay scallop hybrids. The said three-line consists of a male sterile line, a maintainer line and a restorer line. A combination of male sterile line and maintainer line is obtained by continuously backcrossing the male sterile individuals selected from F.sub.1 inter-specific hybrid families with sperm of the selfing family of the sperm-providing bay scallops until the progenies are all male sterile. The restorer line is obtained by continuously backcrossing selected individuals from the male sterile line with sperm of the selfing family of a bay scallop until the progenies are all hermaphroditic and exhibit excellent production traits. Commercial male sterile brood stocks are produced by backcrossing the male sterile line with the maintainer line and commercial hybrid spats are produced by backcrossing the male sterile line and the restorer line.

Claims

1. A three-line breeding system using Peruvian scallop×Bay scallop hybrids, the system comprising: a male sterile line, comprising a Peruvian scallop×Bay scallop hybrid line that is male-sterile and yet female-fertile and spawns only eggs; a maintainer line, comprising a Bay scallop line that provides sperm to fertilize eggs of the male-sterile line to produce a next generation male-sterile line; and a restorer line, comprising a Bay scallop line that provides sperm to fertilize eggs of the male-sterile line to produce commercial hermaphroditic hybrid scallops.

2. A three-line breeding method using Peruvian scallop×Bay scallop hybrids, the method comprising: 1) selecting F.sub.1 male-sterile individuals from F.sub.1 Peruvian scallop×Bay scallop hybrid families and also Bay scallop individuals from the corresponding selfing Bay scallop family, comprising 1a) producing, for step 1b, F.sub.1 hybrid families wherein each family is produced using eggs from a Peruvian scallop and sperm from a Bay scallop, meanwhile each Bay scallop that provides sperm is used to produce a corresponding selfing Bay scallop family for step 1c; 1b) selecting, for step 2a, male-sterile individuals from the F.sub.1 hybrid families produced in step 1a, wherein the individual is determined to be male-sterile if it only spawns eggs; and 1c) selecting, for step 2a, Bay scallop individuals from each corresponding selfing Bay scallop family produced in step 1a; 2) breeding a male-sterile line, comprising: 2a) producing, for step 2b, backcross families wherein each backcross family is produced using eggs from a selected male-sterile F.sub.1 hybrid individual produced in step 1b and sperm from the corresponding selected Bay scallop individual selected in step 1c, meanwhile each Bay scallop that provides sperm is used to produce a corresponding selfing Bay scallop family for step 2b and step 2d; 2b) selecting from the backcross families produced in step 2a, for step 2c, a backcross family wherein each individual in this family is male-sterile/female-fertile, wherein the individual is determined to be male-sterile/female-fertile if it spawns only eggs and the spawned eggs can be fertilized by sperm of the selected Bay scallop individuals from the corresponding selfing Bay scallop family produced in step 2a and produce progenies; 2c) obtaining a male-sterile line, wherein the backcross family produced in step 2b is selected as the male-sterile line; and 2d) selecting, for step 3, the selfing Bay scallop family corresponding to the male-sterile line selected in step 2c; 3) obtaining a maintainer line from step 2d, wherein the selfing Bay scallop family selected in step 2d is selected as the maintainer line specific for the male-sterile line produced in step 2c, as this selfing Bay scallop family is able to render the male-sterile/female-fertile characteristics to the progenies of the male-sterile line produced in step 2c; 4) breeding a restorer line, comprising: 4a) producing, for step 4b, hybrid families using eggs from individuals of the male-sterile line produced in step 2c and sperm of Bay scallops that are not genetically related to the male-sterile line produced in step 2c, meanwhile each Bay scallop that provides sperm is used to produce a corresponding selling family for step 4c; 4b) selecting, from the hybrid families produced in step 4a, a hybrid family wherein each individual in this hybrid family is hermaphroditic, and 4c) selecting a restorer line, which is the selfing Bay scallop family produced in step 4a corresponding to the hybrid family selected in step 4b, as this selfing Bay scallop family is able to restore the male-fertility of the progenies of the male sterile line produced in step 2c; and 5) reproducing the three-line breeding system, wherein a next generation of male-sterile line is produced by fertilizing the eggs of the male-sterile line produced in step 2c with the sperm from the corresponding maintainer line produced in step 3, and the commercial hermaphroditic hybrid scallops are produced by fertilizing the eggs of the male-sterile line produced in step 2c with the sperm from the corresponding restorer line produced in step 4c.

Description

DETAILED DESCRIPTION OF THE EMBODIMENTS

(1) The present invention establishes a three-line breeding system, comprising of a male sterile line, a maintainer line and a restorer line of Peruvian scallop×bay scallop hybrids and also application methods for commercial production of the Peruvian scallop×bay scallop hybrid spats using the three-line breeding system. The specific steps are as follows:

Embodiment 1

(2) 1) Production of the Male Sterile Individuals of the Peruvian Scallop×Bay Scallop Hybrids and Reproduction of the Sperm-Providing Bay Scallops:

(3) In the first year, 50 Peruvian scallops and 50 bay scallops with mature gonads are selected. After air exposure for 30 min., scallops are placed in separate containers pre-filled with 23° C. seawater for spawning induction. After spawning, the eggs and sperm of each scallop are collected separately. Eggs of a Peruvian scallop are mixed with sperm of a bay scallop to produce a Peruvian scallop×bay scallop F.sub.1 hybrid family. A selfing family is produced for each sperm-providing bay scallop by mixing its eggs with its own sperm. In all cases, only a small amount of sperm are used to avoid adverse effect caused by excess sperm. Ideally, each egg should be surrounded by 5-6 sperm. The fertilized eggs are hatched in dark and the resulted D-larvae are collected. At least 20 F.sub.1 hybrid families and the corresponding selfing family of the sperm-providing bay scallop are established. Larval culture, spat nursery and adult grow-out are carried out following the routine culture methods for the Peruvian scallops.

(4) 2) Breeding of the Male Sterile Line of the Peruvian Scallop×Bay Scallop Hybrids:

(5) In the next spring, at least 100 individuals with excellent production traits and apparent male sterile gonad (with no obvious gonadal development in the male part of the gonad) are selected from each F.sub.1 hybrid family. Twenty scallops with excellent production traits are also selected from each selfing families of the corresponding sperm-providing bay scallops. After conditioning, test spawning is carried out to determine if the individuals selected from the F.sub.1 hybrid families spawn only eggs and the eggs can be fertilized and thus to select the male sterile and yet female fertile F.sub.1 hybrid families. Then at least 50 backcross families are constructed with the eggs from selected male sterile/female fertile individuals and sperm from selected individuals from the corresponding selfing families of the sperm-providing bay scallop. The sperm-providing bay scallop brood stocks of the backcross families are again reproduced by self-fertilization. In the following years, the male sterile/female fertile individuals from the backcross families are continuously backcrossed with individuals selected from the selfing family of the sperm-providing Bay scallops, until all the individuals of the backcross progenies exhibit excellent traits and are male sterile and female fertile. The male sterile line of the Peruvian scallop×bay scallop hybrids is thus obtained and the selfing family of the sperm-providing bay scallop becomes the corresponding maintainer line for this male sterile line.

(6) 3) Breeding of the Maintainer Line of the Peruvian Scallop×Bay Scallop Hybrids:

(7) Individuals with excellent production traits are selected from the maintainer line and conditioned to mature. As the scallops of the maintainer line are hermaphroditic animals, the maintainer line is reproduced by self-fertilization.

(8) 4) Breeding of the Restorer Line of the Peruvian Scallop×Bay Scallop Hybrids:

(9) Fifty large male sterile and female fertile individuals are selected from the male sterile line and the maintainer line respectively and conditioned to ripeness. They are then induced to spawn and the eggs from the male sterile individuals are fertilized by the sperm of different bay scallops to establish at least 20 backcross families; meanwhile, the sperm-providing bay scallops are self-fertilized to establish the corresponding self-fertilization families. At harvest, the growth and survival rates of the backcross progenies are measured and the combining ability tests are carried out for each backcross family. In the following year, excellent individuals are selected from both the male sterile line and the corresponding sperm-providing bay scallop selfing family that correlated to the best backcross family with the highest combining ability and highest percentage of hermaphroditic individuals. After conditioning and spawning induction, eggs from the male sterile line are again backcrossed with the sperm of the selected individual from the selfing family of the sperm-providing bay scallop. Same selections are carried each year on the selfing family of the sperm-providing bay scallop until all individuals in the backcross family are hermaphroditic and exhibit excellent production traits. This selfing family of the sperm-providing bay scallop becomes the restorer line for the corresponding male sterile line. The hermaphroditic restorer line is reproduced by self-fertilization and provides sperm for the commercial production of hybrid spats.

(10) 5) Production of Commercial Peruvian Scallop×Bay Scallop Hybrid Spats Using the Three-Line Breeding System:

(11) Each year, the brood stocks of the male sterile line are produced by hybridizing the male sterile line with the sperm of its maintainer line while the brood stocks of the restorer line are reproduced by self-fertilization. In commercial scallop hatcheries, the brood stocks of the male sterile line and the restorer line are selected in the ratio of 5:1 and conditioned to ripeness at the same time. Then the brood stocks of the male sterile line are induced to spawn in large tanks and the brood stocks of the restorer line are induced to spawn in 20 liter buckets. The sperm of the restorer line are collected by filtering the seawater in the buckets through a 500-mesh screen and added to the spawning tanks containing the eggs from the male sterile line brood stocks and well stirred immediately. Seawater samples are taken frequently from the tanks for observation of the fertilization under a microscope until 5-6 sperm are found around each egg. The commercial Peruvian scallop-bay scallop hybrid seeds can thus be produced at large scales and low costs.

Embodiment 2

(12) In late March 2008, mature bay scallops (A. i. irradians) and Peruvian scallops were selected and induced to spawn in separate jars under close observation. The eggs of the Peruvian scallops that did not spawn sperm were collected and fertilized with mixed sperm of different bay scallops to produce F.sub.1 Peruvian scallop×bay scallop hybrid spats. In spring 2009, the scallops of the above F.sub.1 hybrids were conditioned to ripeness and those large scallops with only female part of the gonad developed were selected. The selected F.sub.1 scallops were induced to spawn by air exposure for 30 min followed by thermal shock in jars pre-filled with 23° C. seawater. The eggs of the Peruvian scallop×bay scallop hybrids were then fertilized with sperm of Bay scallops to construct the first-filial generation of backcross cohorts of the Peruvian scallop-bay scallop hybrids (BC.sub.1). In spring 2010, after conditioning in the hatchery, it was found that in the BC.sub.1 cohort, most individuals were hermaphroditic while less than 10% of the individuals were male sterile.

(13) These male sterile scallops can be continuously backcrossed with the sperm of the self-fertilized progenies of the sperm-providing bay scallops. After one to several generations of selective breeding, a combination of a male sterile line (serving as the female parent)-maintainer line (serving as the male parent) can be obtained whose hybridized progenies will be all male sterile and yet female fertile. In commercial operations, the male sterile brood stocks can be produced by fertilizing the eggs from the male sterile line with the sperm from the maintainer line at large scales and low costs.

(14) Once the combination of male sterile line and its maintainer line is established, the restorer line of the male sterile line can be selected in the following way. Large individuals in the male sterile line are selected for conditioning and induced to spawn unfertilized eggs. These eggs are fertilized by the sperm of different, unrelated bay scallops to establish at least 20 backcross families. The families containing highest percentage of hermaphroditic animals and exhibit superior production traits based on combining ability tests are selected. In the following years, the selfing families of the Bay scallops corresponding to the selected backcross families are continuously selected by backcrossing with the male sterile line until all individuals in the resulted backcross families are hermaphrodites and perform excellently in major production traits. The selfing family of the bay scallops then becomes the restorer line for the corresponding male sterile line and can be used together with the male sterile line to produce commercial hybrid spats at large scales and low costs in commercial scallop hatcheries.

Embodiment 3

(15) In spring 2010, the male sterile individuals in the F.sub.1 Peruvian scallop×bay scallop (A. i. irradians) hybrids established in 2009 were selected, conditioned to ripeness and induced to spawn. The eggs from the male sterile individuals in the Peruvian scallop×bay scallop hybrid population were fertilized with sperm of the bay scallops to establish 12 backcross families. In the spring of 2011, it was discovered that, after the scallops were conditioned to ripeness, most individuals in one backcross family were male sterile, suggesting that it is possible to breed the male sterile line by backcrossing the bay scallops with the male sterile individuals selected from the F.sub.1 Peruvian scallop×bay scallop hybrids. In the rest 11 backcross families, most individuals were hermaphroditic, suggesting that it is possible to breed the restorer line by backcrossing the bay scallops with the male sterile individuals selected from the F.sub.1 Peruvian scallop×bay scallop hybrids.

Embodiment 4

(16) In 2012, one (Peruvian scallop×bay scallop)×northern bay scallop backcross families was constructed with eggs from F.sub.1 Peruvian scallop×bay scallop hybrids and sperm from northern bay scallops. At harvest in mid-November 2012, the average shell height, average shell length, average shell width and the average whole weight of the backcross family were increased by 34.1%, 29.8%, 23.0% and 111.5%, respectively, compared with the bay scallops produced and cultured under same conditions and over the same period.

(17) After maturation in the spring of 2013, it was discovered that the progenies of the (Peruvian scallop×bay scallop)×bay scallop were all hermaphrodites, indicating again that the restorer line of the male sterile line can be successfully bred by backcrossing the bay scallops with the male sterile individuals.

Embodiment 5

(18) In 2013 and 2014, 18 F.sub.1 Peruvian scallop×bay scallop (A. i. irradians) hybrid families together with the selfing families of their sperm-providing bay scallops were established. According to the methods given in Embodiment 1, the male sterile families of the Peruvian scallop-bay scallop hybrids and the corresponding maintainer line and restorer line thereof can be bred and the production of commercial Peruvian scallop-bay scallop hybrid spats can be realized within 1-3 years.