Performing Department
(N/A)
Non Technical Summary
Bone weakness and bone damage in laying hens are important economic and animal welfare issues for the poultry industry; skeletal issues in laying hens significantly affect egg production and egg quality. Sclerostin (SOST) is a key regulator of bone, muscle, and fat formation; it reduces bone mineralization, while it inhibits meat production but enhances fat production. However, roles of SOST in regulation of bone, muscle, and fat formationin poultry have not been investigated. Thus, it is a novel approach to enhance bone health, body composition, and feed efficiency in laying hens. The research proposed in this application is significant, because the results from the proposed study will provide new strategies such as genetic selection for inactive SOST or less expression of SOST and development of natural bioactives as feed additives for inaction of SOST to promote bone and muscle development and reduce fat formation while maintaining efficient egg production and improving eggshell quality, increasing economic benefits for the poultry industry.
Animal Health Component
40%
Research Effort Categories
Basic
30%
Applied
40%
Developmental
30%
Goals / Objectives
OVERALL OBJECTIVE: to determine the roles of SOST Knockout (KO) on bone health, muscle development, fat formation, feed efficiency, and egg productin/qaulity in laying hens. CENTRAL HYPOTHESIS: SOST-KO in hens will stimulate bone formation and muscle development and reduce fat formation, improving skeletal health and feed efficiency and maintaining efficient egg production and quality in laying hens. Approach: Generation of SOST KO hens, bone development and health evaluation using microCT, body composition by DEXA, eggshell ultra-structural analyses, feed efficiency, morphological analyses of muscle and fat, and RNAseq. ?
Project Methods
Objective 1: Given the roles of anti-osteogenic, pro-adipogenic, and anti-myogenic activities of SOST, it is hypothesized that gene editing of the SOST gene by our adenoviral CRISPR/Cas9 system will result in pro-osteogenic, anti-adipogenic, and pro-myogenic phenotypes in commercial chicken layers. To generate gene-edited chickens with SOST knockout, several guide RNAs (gRNA) will be selected and evaluated for their efficiencies in inducing gene-edition in vitro. The best gRNAs with high efficiencies will be incorporated into adenoviral CRISPR/Cas9 vectors and the resulting adenoviral vectors will be produced and purified. The purified adenoviral CRISPR/Cas9 vectors will be injected into the subgerminal cavity of chicken blastoderms to induce genome-edition in primordial germ cells. The potential germ-line chimeric hatchling (G0, generation 0) from the injected embryos will be grown to become sexually mature and mated with wild-type chickens. The eggs from the breeding of chimeric chickens and wild-type chickens will be collected and hatched. These G1 offspring (40-60 offsprings per chimeric line) will be screened for gene-edition by sequencing the target area of SOST gene. Male and female G1 heterozygous chickens (SOST+/−) will be bred to generate G2 chickens. The G2 offspring with three different genotypes (SOST+/+, +/−, −/−) will be used for the study described in Objective 2-4. Objective 2: The prevalence of deviation/deformation, fracture, and tip fracture among three groups (SOST+/+, +/−, and −/−) by palpation will be evaluated at 10, 15, 20, 25, 50, 75, and 100 weeks. After that, more detailed keel bone 3-D structural analyses will be performed using microCT. The bone growth and mineralization of femurs (cortical, trabecular and medullary bones) and keel bones of chickens (n=10/group; 0.920-1.000 power) post-hatch ages (10, 15, 20, 25, 50, 75, and 100 weeks for female) will be measured.Objective 3: The daily egg production and first egg laying dates of three groups (SOST+/+, +/−, and −/−) of chickens will be measured from first egg laying dates to 100 weeks. Egg quality (egg weight, yolk weight, albumen weight, egg size, albumen height, haugh unit, yolk color, and eggshell breaking strength) will be measured at 20, 25, 50, 75, and 100 weeks.Objective 4: The body weight of three groups (SOST+/+, +/−, −/−) of female chickens will be measured at 1-week intervals from hatch to 15 weeks before the timing of egg production which causes a significant fluctuation in body weight and egg production significantly affects fat pad weight. Food intake will also be measured to calculate feed efficiency. The adiposity and muscle characteristics of chickens (n=10/group) at embryonic (E18) and post-hatch ages (15 weeks for pre-laying females, and 30 weeks for actively laying hens) will be measured using histomorphometry, DEXA, and RNAseq.