Yu-Shin Cheng

AFLP fingerprinting for paternity testing in ducks

1. The accuracy and reproducibility of AFLP fingerprinting was investigated in the duck (Anas Platyrhynchos), using a multicolour fluorescent labeling technique. The fluorescent labelling fragments were separated on a capillary electrophoresis-base ABI PRISM 3100 Genetic Analyzer. 2. A total of 337 AFLP peaks with 103 of them being polymorphic markers were generated by 16 sets consisting of EcoRI/TaqI primer pair combinations. The number and size range of AFLP polymorphisms detected per primer pair varied from 3 to 11 and 58 to 290 bp, respectively. About 30·6% (103/337) of AFLP peaks were detected polymorphisms, with an average of 6·4 polymorphic markers per primer pair. 3. The clear polymorphic peaks were amplified with EcoR+AC/Taq+AC primer combinations. The AFLP peaks showed high reproducibility. From the family testing, we found that the fingerprints of all the offspring were derived from one or other parent. Therefore, we conclude that AFLP fingerprinting might be a suitable method for duck paternity testing.

Selection responses in duration of fertility and its consequences on hatchability in the intergeneric crossbreeding of ducks.

1. From 1992 to 2003, selected (S) and control lines (C) of the laying Brown Tsaiya duck (Anas platyrhynchos) were simultaneously maintained under the same standardised conditions of feeding and management. 2. The selection objective was to increase the number of fertile eggs after a single artificial insemination (AI) with pooled Muscovy semen. From generations G1 to G11, 2452 and 2022 female ducks, in S and C lines, respectively, were measured and recorded. In the S line, the percentage selected varied between 20·2 and 34·3% in females and between 7·2 and 20·8% in males. 3. Selection for number of fertile eggs had a correlated effect of increasing the parameter τ of the logistic curves which fitted the daily variations (d 2 to 15) in fertility or hatchability on the basis of eggs set. The differences S − C for the estimates of the times of half maximal fertility and hatchability increased by 0·41 and 0·37 d per generation between G1 and G11, respectively. 4. The highest increases of fertility per day rates after a single AI were observed between d 5 and 11. Moreover, in the selected line, fertility rate was higher than, or equal to, 90% in d 2 from G8. The same tendencies were observed for the changes in the evolution of hatchability on the basis of eggs set. 5. Selection increased fertility and hatchability according to the egg set rates, especially for d 2 to 8 after AI. Hatchability of fertile eggs was not impaired, confirming that selection for one AI per week was possible in this strain of laying ducks.

A novel genetic marker of the ovomucoid gene associated with hatchability in Tsaiya ducks (Anas platyrhynchos)

Transcriptome analysis using a cDNA microarray was performed to identify differentially expressed genes that are correlated with hatchability, and a new PCR-RFLP marker of high hatchability among the identified genes was observed. We used the cDNA microarray technique for gene expression profiling of the magnum epithelium of laying Tsaiya ducks, and several regulated genes associated with hatchability were found. The results of real-time PCR and Western blotting analysis confirmed that the mRNA and protein levels of ovomucoid in the magnum epithelium of animals in the low-hatchability group were significantly higher than the levels in the high-hatchability group (P < 0.05). Primers TovF1 and TovR1, designed according to the ovomucoid EST sequence, were used to amplify genomic DNA samples of different individual Tsaiya ducks, and sequence analysis of the amplified DNA products showed deletion among the ducks from the low-hatchability group. Primers TovF2 and TovR2 were used to perform PCR-RFLP analysis on the amplified DNA products to classify the ducks into +/+, +/- and -/- genotypes. The animals of +/+ and +/- genotypes were identified as having significantly higher hatchability than those of the -/- genotype (P < 0.05). In contrast, no differences were observed between genotypes in terms of fertility, duration of fertility, egg weight or total number of eggs. Our results indicated that a novel PCR-RFLP marker of high hatchability, an ovomucoid gene polymorphism, can be used as a genetic marker for marker-assisted selection to improve hatchability in Tsaiya ducks.

Duck (Anas platyrhynchos) linkage mapping by AFLP fingerprinting

Amplified fragment length polymorphism (AFLP) with multicolored fluorescent molecular markers was used to analyze duck (Anas platyrhynchos) genomic DNA and to construct the first AFLP genetic linkage map. These markers were developed and genotyped in 766 F2 individuals from six families from a cross between two different selected duck lines, brown Tsaiya and Pekin. Two hundred and ninety-six polymorphic bands (64% of all bands) were detected using 18 pairs of fluorescent Taq I/Eco RI primer combinations. Each primer set produced a range of 7 to 29 fragments in the reactions, and generated on average 16.4 polymorphic bands. The AFLP linkage map included 260 co-dominant markers distributed in 32 linkage groups. Twenty-one co-dominant markers were not linked with any other marker. Each linkage group contained three to 63 molecular markers and their size ranged between 19.0 cM and 171.9 cM. This AFLP linkage map provides important information for establishing a duck chromosome map, for mapping quantitative trait loci (QTL mapping) and for breeding applications.

Factors Affecting the Incidence of Angel Wing in White Roman Geese: Stocking Density and Genetic Selection

The present study investigated stocking density and genetic lines, factors that may alter the severity and incidence of angel wing (AW), in White Roman geese. Geese (n = 384) from two genetically selected lines (normal- winged line, NL, and angel-winged line, AL, respectively) and one commercial line (CL) were raised in four pens. Following common commercial practice, low-stocking-density (LD), medium-stocking-density, and high-stocking-density treatments were respectively administered to 24, 32, and 40 geese per pen at 0 to 3 weeks (1.92 m2/pen) and 4 to 6 weeks (13.2 m2/pen) of age and to 24, 30, and 36 geese at 7 to 14 weeks (20.0 m2/pen) of age. The results revealed that stocking density mainly affected body weight gain in geese younger than 4 weeks, and that geese subjected to LD had a high body weight at 2 weeks of age. However, the effect of stocking density on the severity score of AW (SSAW) and incidence of AW (IAW) did not differ significantly among the treatments. Differences were observed among the genetic stocks; that is, SSAW and IAW were significantly higher in AL than in NL and CL. Genetic selection generally aggravates AW, complicating its elimination. To effectively reduce IAW, stocking density, a suspected causal factor, should be lower than that presently applied commercially.

A new method for detection of single nucleotide polymorphism in a female reproduction-associated gene, tmigd1, of Anas platyrhynchos using a strip biosensor with gold nanoparticles

In this study, we first reported a lateral flow assay combined with primer extension (PEXT) and gold nanoparticles for single-nucleotide polymorphism (SNP) genotyping of the tmigd1 gene of the Tsaiya ducks (Anas platyrhynchos), which has the advantages of simplicity of operation, cost-effectiveness, and time-saving. Gold nanoparticles were tailed with thiol-thymine oligodeoxyribonucleotides (thiol-(dT)30) using the salt-aging method at 25°C and used as a label in a lateral flow assay. The lateral flow device was composed of test and control zones on a nitrocellulose membrane containing streptavidin and adenosine oligodeoxyribonucleotides ((dA)30), respectively. When the specific SNP existed, the corresponding primers were extended, and the reaction product was captured by streptavidin at the test zone owing to the introduction of biotin-deoxyuridine triphosphate (biotin-dUTP) into the reaction product during PEXT. Gold nanoparticles hybridized with the reaction product to render it visible. Here, we developed a new system for detection of single nucleotide polymorphism in a female reproduction-associated gene, tmigd1, of Anas platyrhynchos using the strip biosensor, and identified the optimized parameters for the concentration of Mg2+ in the PEXT reaction and the amount of streptavidin used on membranes for signal specificity.

Selection for Duration of Fertility and Mule Duck White Plumage Colour in a Synthetic Strain of Ducks (Anas platyrhynchos)

A synthetic strain of ducks (Anas platyrhynchos) was developed by introducing genes for long duration of fertility to be used as mother of mule ducklings and a seven-generation selection experiment was conducted to increase the number of fertile eggs after a single artificial insemination (AI) with pooled Muscovy semen. Reciprocal crossbreeding between Brown Tsaiya LRI-2 (with long duration of fertility) and Pekin L-201 (with white plumage mule ducklings) ducks produced the G0. Then G1 were intercrossed to produce G2 and so on for the following generations. Each female duck was inseminated 3 times, at 26, 29, and 32 weeks of age. The eggs were collected for 14 days from day 2 after AI. Individual data regarding the number of incubated eggs (Ie), the number of fertile eggs at candling at day 7 of incubation (F), the total number of dead embryos (M), the maximum duration of fertility (Dm) and the number of hatched mule ducklings (H) with plumage colour were recorded. The selection criterion was the breeding values of the best linear unbiased prediction animal model for F. The results show high percentage of exhibited heterosis in G2 for traits to improve (19.1% for F and 12.9% for H); F with a value of 5.92 (vs 3.74 in the Pekin L-201) was improved in the G2. Heritabilities were found to be low for Ie (h2 = 0.07±0.03) and M (h2 = 0.07±0.01), moderately low for Dm (h2 = 0.13±0.02), of medium values for H (h2 = 0.20±0.03) and F (h2 = 0.23±0.03). High and favourable genetic correlations existed between F and Dm (rg = 0.93), between F and H (rg = 0.97) and between Dm and H (rg = 0.90). The selection experiment showed a positive trend for phenotypic values of F (6.38 fertile eggs in G10 of synthetic strain vs 5.59 eggs in G4, and 3.74 eggs in Pekin L-201), with correlated response for increasing H (5.73 ducklings in G10 vs 4.86 in G4, and 3.09 ducklings in Pekin L-201) and maximum duration of the fertile period without increasing the embryo mortality rate. The average predicted genetic response for F was 40% of genetic standard deviation per generation of selection. The mule ducklings’ feather colour also was improved. It was concluded that this study provided results for a better understanding of the genetics of the duration of fertility traits in the common female duck bred for mule and that the selection of a synthetic strain was effective method of improvement.