Sea Hwan Sohn

Production of germline transgenic chickens expressing enhanced green fluorescent protein using a MoMLV-based retrovirus vector

The Moloney murine leukemia virus (MoMLV) ‐based retrovirus vector system has been used most often in gene transfer work, but has been known to cause silencing of the imported gene in transgenic animals. In the present study, using a MoMLV‐based retrovirus vector, we successfully generated a new transgenic chicken line expressing high levels of enhanced green fluorescent protein (eGFP). The level of eGFP expression was conserved after germline transmission and as much as 100 g of eGFP could be detected per 1 mg of tissue protein. DNA sequencing showed that the transgene had been integrated at chromosome 26 of the G1 and G2 generation transgenic chickens. Owing to the stable integration of the transgene, it is now feasible to produce G3 generation of homozygous eGFP transgenic chickens that will provide 100% transgenic eggs. These results will help establish a useful transgenic chicken model system for studies of embryonic development and for efficient production of transgenic chickens as bioreactors.—Koo, B. C., Kwon, M. S., Choi, B. R., Kim, J‐H., Cho, S‐K., Sohn, S. H., Cho, E. J., Lee, H. T., Chang, W., Jeon, I., Park, J‐K., Park, J. B., Kim, T. Production of germline transgenic chickens expressing enhanced green fluorescent protein using a MoMLV‐based retrovirus vector. FASEB J. 20, 2251–2260 (2006)

The Effects of Dietary Supplementation of Vitamin C and E on the Growth Performance and the Stress Response in Broiler Chickens

Department of Animal Science and Biotechnology, Gyeongnam National University of Science and Technology, Jinju 660-758, KoreaABSTRACT This study was performed to investigate the investigated effects of dietary supplementation of vitamin C and E on the growth performance and stress response in broiler chickens. Stress response was analyzed by the quantity of telomeric DNA, the rate of DNA damage and the expression levels of heat shock proteins (HSPs) and hydroxyl-3-methyl-glutaryl coenzyme A reductase (HMGCR) genes on tissues and blood. The telomere length and telomere shortening rates were analyzed by quantitative fluorescence in situ hybridization on the nuclei of lymphocytes and tissues. The DNA damage rate of lymphocytes was quantified by the comet assay. The expression levels of HSP70, HSP90s and HMGCR genes were measured by quantitative real-time polymerase chain reaction in lymphocytes. In results, there was no significant difference among treatments in body weight, weight gain, feed intake and mortality. The telomere shortening rate of the lymphocytes was significantly lower in the vitamin E supplemented group than the control group. The DNA damage was also decreased supplemented with vitamin C and E, as compared to the control group. The vitamin E supplemented group had a significant positive effect on the expressions of HMGCR, HSP90-α and HSP90-β in lymphocytes, but had no significance on HSP70, as compared to the control group. We concluded that the dietary supplementation of vitamin E (100 mg/kg feed) had reduced the individual physiological stress response without stunt growth in broiler chickens.(Key words : vitamin C, vitamin E, growth performance, stress, broiler)

Comparison of Stress Response between Korean Native Chickens and Single Comb White Leghorns subjected to a High Stocking Density

Department of Animal Science and Biotechnology, Gyeongnam National University of Science and Technology, Jinju 660-758, KoreaABSTRACT With Single Comb White Leghorn (WL) and Korean Native Chicken (KNC) breeds, we compared the stress response with chicken breeds that were subjected to a high stocking density. Stress response was analyzed by the quantity of telomeric DNA, the rate of DNA damage and the expression levels of heat shock proteins (HSPs) and hydroxyl-3- methyl-glutaryl coenzyme A reductase (HMGCR) genes on tissues and blood. The telomere length and telomere shortening rates were analyzed by quantitative fluorescence in situ hybridization on the nuclei of lymphocytes and tissues. The DNA damage rate of lymphocytes was quantified by the comet assay. The expression levels of HSP70, HSP90-α, HSP90-β and HMGCR genes were measured by quantitative real-time polymerase chain reaction in lymphocytes. There was no significant difference between KNC and WL in body weight, weight gain, telomere shortening rate and DNA damage rate. However, the growth rate significantly decreased in chickens raised under high stocking density conditions, as compared to the control group. The telomere-shortening rate, DNA damage and HSPs expression of the lymphocytes were significantly higher in the high stocking density group than the control. The stress condition and breeds had a significant effect on the expressions of HSP70, HSP90-α and HSP90-β in lymphocytes, except HMGCR. The stress response of WL was higher than that of KNC, as analyzed to the expression of HSP70 and HSP90-α. Therefore, we concluded that the chickens which were exposed to a high stocking density had increased the individual physiological stress response regardless of breeds, and White Leghorns are more susceptible to stress condition than Korean Native Chickens.(Key words : stress response, telomere, DNA damage, HSP, Korean Native Chicken, Leghorn)

Amount of Telomeric DNA on Pig Lymphocytes by Quantitative Fluorescence in situ Hybridization

ABSTRACT The amount of telomeric DNA was quantified across different breeds (Landrace, Duroc, Yorkshire and Berksire), at different ages (90 days old and 180 days old) and among sexes (male and female) in pigs raised at the Performance Testing Station of Korea Swine Association, Jinkyo, Korea. The telomeric DNA amount was quantified by Quantitative Fluorescence In Situ Hybridization (Q-FISH) using a porcine telomeric DNA probe on interphase nuclei of lymphocytes. Analysis revealed that the amount of telomeric DNA on the pig lymphocytes was found to decrease with age. The quantity of telomeres significantly differed among breeds at 90 days of age. The colored breeds such as Berkshire and Duroc had higher amount of telomeric DNA than the Yorkshire and Landrace breed. In addition, the amount of telomeric DNA in male lymphocytes was significantly higher than that of females. In the correlation coefficients between the telomere quantity and their productive traits; average daily gain, loin percent and index value were positively correlated, whereas body length, feed efficiency and back fat thickness correlated negatively. However, the correlation coefficients were very low and not significant. Therefore, this study suggests that the amount of telomeres on lymphocytes can be considered as a physiological marker but not as a productive marker in pig.(

Analysis of Stress Response of Domestic Chicken Breeds for the Development of a New Synthetic Parent Stock

We compared the degrees of stress response of 12 domestic purebred chicken strains that have been bred at National Institute of Animal Science, RDA, Korea since 1980. As a physiological marker of stress response, the expression levels of heat shock protein (HSP)-70, HSP-, HSP-, hydroxyl-3-methyl-glutaryl coenzyme A reductase (HMGCR) genes and telomere length were measured by quantitative real-time polymerase chain reaction using the lymphocytes of 1,101 chickens. There was significant difference in HSP-70, HSP-, HMGCR expression and telomere length among the strains. There was also significant difference in HSP-, HSP-, and HMGCR expression between male and female chickens. Different age groups of chicken exhibited different expression levels of HSP-70, HSP- and telomere length. The results of the HSPs expression level suggested that, the strains of R, L and Y were highly resistant to stress, whereas the strains of S, O and W were susceptible to stress. Although the statistical differences in some of HSPs gene expression existed between genders, the HSP expression results varied in different strains that some opposed to the others, and there might be interaction between strains and genders, which conclude that there was no difference in stress response between male and female chickens. Moreover, despite of significant difference in some of HSPs expression level, it was considered that there was no difference in stress response between ages due to the inconsistent trends among HSP markers.

The Effects of Dietary Supplementation of Vitamin C or E on the Expressions of Endoplasmic Reticulum Stress, Lipid and Glucose Metabolism Associated Genes in Broiler Chickens

This study was conducted to evaluate the effects of dietary supplementation of vitamin C or E on the expressions of endoplasmic reticulum (ER) stress, lipid and glucose metabolism associated genes in broiler chickens. A total of 216 one-day-old male broilers was randomly alloted to 4 treatments with 6 replicate pens per treatment and 9 broilers per pen for 35 days. The dietary treatments were control, vitamin C (control diet + ascorbic acid 200 mg/kg diet), vitamin E (control diet + -tocopherol 100 mg/kg diet), vitamin C + E (control diet + vitamin C 200 mg/kg + vitamin E 100 mg/kg), respectively. To evaluate gene expressions by quantitative real-time polymerase chain reaction, total RNA was extracted from the liver of the chicken at 35 days of age. Dietary supplementation of vitamins was significantly down-regulated the expression of stress marker genes including HSP70, HSP90, and HMGCR, as compared to the control (p

Genotype Frequencies of the Sex-Linked Feathering and Their Phenotypes in Domestic Chicken Breeds for the Establishment of Auto-Sexing Strains

The method of sexing based on differences in the rate of feather growth provides a convenient and inexpensive approach. The locus of feather development gene (K) is located on the Z chromosome and can be utilized to produce phenotypes that distinguish between the sexes of chicks at hatching. To establish the auto-sexing native chicken strains, this study analyzed the genotype frequency of the feathering in domestic chicken breeds. The method of classification of slow- and rapid-feathering chickens was also investigated. In the slow-feathering chicks, the coverts were either the same length or longer than the primary wing feathers at hatching. However, the rapid-feathering chicks had the primary wing feathers that were longer than the coverts. The growth pattern of tail feather also distinctively differed between the rapid- and slow-feathering chicks after 5-days. The accuracy of wing feather sexing was about 98% compared with tail sexing. In domestic chicken breeds, Korean Black Cornish, Korean Rhode Island Red, and Korean Native Chicken-Red had both dominant (K) and recessive () feathering genes. The other breeds of chickens, Korean Brown Cornish, Ogol, White Leghorn, Korean Native Chicken-Yellow, -Gray, -White and -Black had only the recessive feathering gene (). Consequently, feather sexing is available using the domestic chicken breeds. Establishing the maternal stock with dominant gene (K-) and paternal stock with recessive gene (), the slow-feathering characteristic is passed from mothers to their sons, and the rapid-feathering characteristic is inherited by daughters from their fathers.

The Effect of Storage Period and Temperature on Egg Quality in Commercial Eggs

Consumers demand fresh and high-quality eggs. Egg quality may be represented by shell color, shell weight, egg weight, shell thickness, shell density, albumen height, yolk color, albumen pH and viscosity. Various factors such as strain, age of hen, storage temperature, humidity, the presence of CO² and storage time affect egg quality. Therefore, we investigated the effects of storage time and temperature on egg quality to define the freshness of Korean market eggs. A total of 1,800 eggs were used for this experiment and were separated into 45 treatments with 40 eggs in each. The treatments were consisted of 15 storage periods (2 d to 30 d) and 3 storage temperatures (2℃, 12℃, 25℃). Each egg was weighed and broken, and the height of the thick albumen, Haugh units (HU), egg shell color and yolk color were measured by a QCM+system. We also observed the physiochemical properties of eggs such as yolk pH, albumen pH and albumen viscosity. The egg weight, shell weight, albumen height, HU and albumen viscosity significantly decreased with increasing storage time and temperature. However, the albumen and yolk pH significantly increased with increasing storage period and temperature. The interaction effects between the storage period and temperature were significant for shell weight, shell density, egg weight, albumen height, HU, yolk color, yolk pH, albumen pH and albumen viscosity. In the analysis of the correlation with egg quality, the storage temperature exhibited a higher correlation coefficient than the storage period. In conclusion, storage time and temperature are the major factors affecting egg quality, but the storage temperature is a more sensitive determinant of egg quality deterioration compared with the storage period.

Effects of High Stocking Density on the Expression of Metabolic Related Genes in Two Strains of Chickens

Chickens are exposed to the external and internal stressors such as low and high temperature, high stocking density, feed restriction and disease. There have been a few studies on gene expressions through the investigation of chickens under direct exposure to the stress of high stocking density. The objective of the present study was to determine the expressions of genes associated with stress, endoplasmic reticulum (ER)-stress, lipid and glucose metabolism in two strains of chickens, Korean Native Chicken (KNC) and White Leghorn (WL), raised in high stocking density. A total of 164 chickens aged 40 weeks were randomly allotted to a 540 cm 2 /bird stocking density (control), whereas the chickens in a high density group were assigned in a 311 cm 2 /bird stocking density with feeding ad libitum for 10 weeks. Total RNA was extracted from the live for qRT-PCR. The expression levels of hsp70 and hsp90α were higher in WL subjected to stress with high stocking density compared with those genes in control (P<0.05), while the expressions of genes were not affected in KNC. ER stress marker gene XBP1 was also highly expressed in WL with stress (P<0.05), but the stress of high stocking density did not influence to ER stress marker genes in KNC. Lipid metabolism associated genes including FABP4, FATP1 and ACSL1 were highly expressed in WL compared with KNC when subjected to high stocking density stress (P<0.05). The expression of glucose transport gene GLUT2 and GLUT8 were increased in chickens exposured to the stress of high stocking density (P<0.05). The data indicate that WL is more sensitive to the stress of high stocking density compared with KNC and the stress may influence the modulation of lipid and glucose metabolism in the liver of chickens.

Influence of Early- and Late-feathering Phenotype on Productive Performance in the Feather-sexing Strains of Korean Native Chicken

The vent sexing and the auto-sexing by using sex-linked traits are general sexing methods of day-old chicks. Currently, the feather sexing which is based on the differences in the feather characteristics at hatching is the representative sexing method of chicken, because the late-feathering is sex-linked trait. The feather sexing can be used if the breed has dominant feathering gene (K) in maternal and recessive gene (k + ) in paternal. Therefore it is necessary to identify the associa- tion of feathering genes and quantitative traits in chickens. In this study, we investigated the influence of the rate of feathering on productive traits in Korean Native Chicken. In results, there was no significant difference between early-feathering chickens and late-feathering chickens in reproductive performance such as fertility and hatchability. Livability, body weights, egg production, egg weight and egg quality also did not significantly differ between early- and late-feathering chickens. Age at first egg was the only trait of those tested in which significant difference was observed. The early-feathering chickens laid eggs 3 days earlier than late-feathering chicken. As a result, there is no influence of feathering phenotypes on productive performance in Korean Native Chickens. Consequentially, establishing the feather sexing strain is available using the Korean Native Chicken breed without considering of the effect of feathering genes on productive traits.