A practical approach was proposed to produce transgenic chimeric chickens using blastodermal cells (BCs). The chicken BCs were mechanically dissociated and transferred into the recipient eggs that had been exposed to 500 rads irradiation of$^{60}Co$ and windowed on the equatorial plane. Chimeric chickens were generated using two models: the crosses (MXL) from Black Minors (ii,EE,b/b) ♂${\times}$Barred Leghorns (ii,ee,B/-) ♀ as donors and White Leghorns (WL, II) as acceptors (Model 1), or the Black Heifengs (BH, ii,EE,bb) as donors and Hua-xing white (HW, II) as recipients (Model 2). The treated eggs were incubated in their original shells in normal conditions until hatching. Green fluorescent protein (GFP) gene was transferred into the BCs derived from MXL and BH via lipofectamine and the pEGFP-C1, and transfection efficiency into the BCs was examined under a fluorescent microscope. Potential transgenic chimeras were selected based on the proposed methods in this study. Using the fresh BCs, the best rate of phenotypic chimeras was 6.7% and 26.0% in model-1 groups, and model-2 groups, respectively. We also described the optimized conditions for transfection. Although 30% of the BCs transfected in vitro emitted green light under an inverted fluorescent microscope, no embryos injected with the transfected BCs expressed foreign GFP gene at 3-4 days.
The objective of this study was to establish conditions for transfection of a foreign gene into somatic cells using cationic lipid reagents and to evaluate the effects of transfection on in vitro development of somatic cell nuclear transfer (SCNT) embryos. Green fluorescent protein (GFP) gene was used as a foreign gene and a non-transfected somatic cell was utilized as a control karyoplast. Monolayers of porcine cells were established and subsequently transfected with a GFP-expressing gene (pEGFP-N1) using three types of transfection reagents (LipofectAMINE PLUS, FuGENE 6 or ExGen500). Donor cells used for SCNT included transfected fetal or adult fibroblasts and oviduct epithelial cells, either serum-fed or serum-starved. Oocytes matured in vitro for 42 h were reconstructed with either transfected or non-transfected porcine somatic cells by electric fusion and activation using a single DC pulse of 1.8 kV/cm for $30{\mu}s$ in $Ca^{2+}$ and $Mg^{2+}-containing$ 0.26 M mannitol solution. Reconstructed oocytes were subsequently cultured in NCSU-23 medium for 168 h and the developmental competence and cell number in blastocyst were compared. There were no significant differences (P>0.05) in fusion, cleavage rates or development to the blastocyst stage between non-transfected, transfected, serum-fed and serum-starved cells. However, the rates of GFP-expressing blastocysts were higher in the FuGENE 6 group (71.4%) among transfection reagents and in the fetal fibroblasts group (70.4%) for donor cells. These results indicate that fetal fibroblasts transfected with FuGENE 6 can be used as donor cells for porcine SCNT and that GFP gene can be safely used as a marker of foreign genes in porcine transgenesis.
Hwang, Eun-Sun;Duncan, Claudine E.;Bowen, Phyllis E.
Preventive Nutrition and Food Science
/
v.14
no.4
/
pp.343-348
/
2009
This study sought to determine changes in self-selected diets in response to a high energy tomato pasta entree. Thirty men, mostly African-American, who were diagnosed with prostate cancer and also scheduled for prostatectomy were enrolled in the study. Dietary intakes were obtained by 24 hr diet recall for 3 days before the intervention and 3 days in week 2 during the intervention. Tomato pasta entrees were formulated to contain 30 mg of lycopene with roughly the same macronutrient composition and averaged 771 kcal/entree. Mean adherence to lycopene dose was 82% and, days when the entree was consumed, the mean adherence to lycopen was 90%. Lycopene intake in their self-selected diet decreased from 5 to 1 mg/day which lowered the exposure to lycopene, planned during the intervention. The men were able to decompensate for most of the added energy to their diet, but there was still a mean increase in total intake of 242 kcal/day (p=0.04), which did not result in weight gain over the short period of the study. There were no significant changes in the percent of energy from protein, fat or carbohydrates, but dietary cholesterol increased from 341 to 472 mg/day (p=0.0002). Mean intakes of vitamins A and C, and folate were not significantly different and were above the EAR. Vitamin E intake decreased, but, because of the variance in intake, the decrease was not statistically significant. Possible deterioration of diet quality should be a consideration when recommending whole food interventions for the prevention of chronic disease or the amelioration of physiological dysfunction.
This experiment was planned to develop a functional supplements by food resources to prevent and lessen the bad effects caused by the environmental pollution such as bad food, air, water and heavy metals exposed to people these days. As for Its primary stage, the nutrient profile and sensory characteristics of the duck-extract were evaluated in this study. The duck-extract was formulated by the mixture of bone and internal organs of ducks and 6 medicinal herbs which were able to be used as foods and known to help the excretion and the repression of the poison inside the body for a long time. As we compared the six medicinal herbs mentioned above to the vegetable herbs, the nutrient profile of the medicinal herbs were superior to the vegetable herbs. The duck-extract was composed of protein 49.92%, carbohydrate 37.02%, dietary fiber 20.99%, lipid 7.60% and ash 5.46%. The ratio of Ca to P was 1 : 1.4, which was a suitable ratio for absorption, the contents of Na and K were low and those of micro-elements such as Fe, Mn, Zn and Cu high compared to other meat extracts. The essential amino acids accounted for 30.91% of total amino acids. The result of sensory evaluation was better in overall preferences than the ones which are already in the market. From these results, the nutrient profile of the duck-extract was estimated to be able to supply enough nutrients to the people whose nutritional balance was destroyed these days. This study also showed the effective way of using duck-extract and its application to the oriental medicine.
Objectives: Expression of vascular endothelial growth factor C (VEGF-C)and vascular endothelial growth factor feceptor-3 (VEGFR-3) in laryngeal squamous carcinoma and its relationship to lymph node metastasis were investigated. Methods: VEGF-C and VEGFR-3 gene expression in 30 cases of normal laryngeal mucosa tissue (NLM), primary laryngeal carcinoma cell carcinomas (PLC) and cervical lymph nodes (CLN) was examined by reverse transcription polymerase chain reaction (RT-PCR). Protein levels of VEGF-C expression were determined by immunohistochemical staining in 60 cases of PLC. Results: Expression of VEGF-C and VEGFR-3 different among NLM, PLC and CLN in the same patient. In PLC, expression was significantly higher in lymph node positive group than in the lymph node negative group and associated with histological grade of differentiation; Expression of VEGF-C and VEGFR-3 was not linked with age, sex, site or T stage. Conclusions: A close correlation was found between VEGF-C/VEGFR-3 expression and lymph node metastasis in PLC, suggesting a role in metastasis of laryngeal carcinomas.
Journal of the Korean Society of Food Science and Nutrition
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v.27
no.6
/
pp.1034-1040
/
1998
The nutritional quality and physiological activity of cultivated hoelen from 13 months to 24 months were examined to compare with natural hoelen. General compositions of hoelen were as follows re spectively; crude protein 1.80~2.50%, crude fat 0.68~1.23%, crude ash 0.12~0.43%, crude fiber 6.30~7.14%, nitrogen free extract 89.25~90.44%. The major free sugar was glucose, and the content of 13 months cultivated hoelen was higher than any other cultivated hoelen. The major fatty acids of hoelen were linoleic acid, palmitic acid, stearic acid and oleic acid. The ratio of unsaturated fatty acids was 62.62~77.96% and the content was higher in cultivated hoelen than in natural hoelen. The contents of amino acid were high in the order of tyrosine, aspartic acid, glutamic acid and serine, and increased as the day of cultivation became longer. The contents of mineral components were higher in the order of Ca, K, Mg and the contents of Na and Ca were higher in natural hoelen than in cultivated hoelen. The contents of pachymic acid and dehydropachymic acid was similar in the natural and cultivated hoelen. On the results of antimicrobial test the ethanol extract showed a stronger effect than water extract, and had an excellent antimicrobial activity on Micrococcus luteus and Escherichia coli. The ethanol extract of hoelen showed comparatively strong electron donating ability. The ethanol extract of hoelen showed a high inhibition activity on the growth of lung cancer, ovary cancer, skin cancer, central nerve cancer and rectum cancer, especially the activity of 19 months cultivated hoelen was the highest. On the above results of nutritional quality and physiological activity of hoelen, it is supposed that the picking date of cultivated hoelen was suitable over than 19 months.
Shim, Sang Woo;Kim, Young Ha;Lee, Hoon Taek;Shim, Hosup
Asian-Australasian Journal of Animal Sciences
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v.21
no.3
/
pp.358-363
/
2008
The intracytoplasmic sperm injection (ICSI) procedure has recently been utilized to produce transgenic animals and may serve as an alternative to the conventional pronuclear microinjection in species such as pigs whose ooplasm is opaque and pronuclei are often invisible. In this study, the effects of sperm membrane disruption and electrical activation of oocytes on in vitro development and expression of transgene green fluorescent protein (GFP) in ICSI embryos were tested to refine this recently developed procedure. Prior to ICSI, sperm heads were treated with Triton X-100+NaCl or Triton X-100+NaCl+NaOH, to disrupt membrane to be permeable to exogenous DNA, and incubated with linearized pEGFP-N1 vector. To induce activation of oocytes, a single DC pulse of 1.3 kV/cm was applied to oocytes for $30{\mu}sec$. After ICSI was performed with the aid of a micromanipulator, in vitro development of embryos and GFP expression were monitored. The chemical treatment to disrupt sperm membrane did not affect the developmental competence of embryos. 40 to 60% of oocytes were cleaved after injection of sperm heads with disrupted membrane, whereas 48.6% (34/70) were cleaved without chemical treatment. Regardless of electrical stimulation to induce activation, oocytes were cleaved after ICSI, reflecting that, despite sperm membrane disruption, the perinuclear soluble sperm factor known to mediate oocyte activation remained intact. After development to the 4-cell stage, 11.8 (2/17, Triton X-100+NaCl+NaOH) to 58.8% (10/17, Triton X-100+NaCl) of embryos expressed GFP. The expression of GFP beyond the stage of embryonic genome activation (4-cell stage in the pig) indicates that the exogenous DNA might have been integrated into the porcine genome. When sperm heads were co-incubated with exogenous DNA following the treatment of Triton X-100+NaCl, GFP expression was observed in high percentage (58.8%) of embryos, suggesting that transgenic pigs may efficiently be produced using ICSI.
The objectives of this study were to measure particle size and evaluate the effect of increasing alfalfa hay particle size on production characteristics in lactating Holstein dairy cows. Ninety multiparous Holstein cows in early to mid-lactation were randomly assigned in a complete randomized design for a 30-day period. Animals were offered one of the three diets, which were identical in energy, protein, and chemical composition, but differed only in particle size of alfalfa hay. The treatments were A) total mixed ration (TMR) in which only fine chopped alfalfa hay was incorporated in the ration, B) the same diet in which half of the alfalfa hay was fine chopped and incorporated in the mixed ration and half was long hay and offered as a top dressing, and C) the same diet with long hay alfalfa offered as a top dressing. Distribution of particle size of rations was determined through 20,000, 8,000 and 1,000 ${\mu}m$ sieves. The new method of quantitative determination of manure index was examined for each cow on different treatments. The geometric mean length of particle size in the rations was 5,666, 9,900 and 11,549 ${\mu}m$ for treatments A, B and C, respectively. Fat corrected milk (4%), milk fat percentage and production were significantly different (p<0.05) in treatment A versus B and C (fat corrected milk (FCM, 4%)) 28.3 vs. 35.2 and 32.3 kg/d, fat percentage 2.89, 4.04 and 3.62; but the change of ration particle size had no significant effect on milk production (p>0.05). Blood concentration of cholesterol in treatment A was significantly higher (p<0.05) than treatment B and C (181.0 vs. 150.0 and 155.2 mg/dl). Manure index in treatment C was significantly different (p<0.05) from treatment B (15.86 vs. 17.67). Based on these experimental findings, it is concluded that an increase in the ration particle size can increase milk fat percentage due to providing more physically effective fiber, which in turn could effect changes in manure consistency.
In order to investigate if the employment of housewives may affect the nutritional status of their family members, an analysis was made for the data of 2001 Korea National Health and Nutrition survey. Housewives aged 20 or over were divided into two groups of the working (W, 44.3%) and the non-working (NW, 55.7%), and household income levels were divided into 4 groups of low, middle, high, and high above according to the minimum cost of living in the year of 2001. Nutrient intakes were assessed by using dietary recommended intakes for Koreans of 2005. Working housewives showed similar levels to those of non-working housewives in most nutrients intakes except energy and vitamin C. However their families excluding housewives of W, than those of NW, took less protein, calcium, iron, potassium, vitamin A, thiamin, riboflavin, niacin, and vitamin C when assessed as % of recommended intakes and took more sodium. Such differences were very strong in children and adolescents, and in the middle income households. More % of the families of W than those of NW consumed nutrients below the estimated average requirements. Percents of hypertension classified by both systolic and diastolic blood pressure were higher in adult family members of W than in those of NW. This tendency seemed to be more significant in the family members aged 30 to 49. Both obesity and under-weight rates of school children ($7{\sim}12\;yrs$) in W were higher than those in NW. The above resuIts suggested that employment of housewives could have negative influences on the nutritional status of their family members, especially of their children and in the middle income class.
Background: Liquid feeding system has been introduced to domestic swine farms, but negative cognition about liquid feeding system has been remained for feed waste decay related with poor management and microbial contamination. For these reasons, this study was conducted to evaluate the effects of feeding method in lactating sows. Methods: A total of 30 mixed-parity (average 4.13) lactating sows (Yorkshire ${\times}$ Landrace) with an initial BW of $218.8{\pm}19.5kg$ was used in a 3 week trial. Sows were allotted to 1 of 2 treatments in a completely randomized design by their body weight, backfat thickness, parity and alive litter weight. One of treatments was dry feeding and the other was liquid feeding (water to feed ratio, 1:1). Experimental diets contained 3265 kcal ME/kg, 12.6 % CP, 5.76 % EE, 1.09 % total lysine, 0.25 % total methionine, as fed basis. Results: Dry feeding treatment had high body weight loss rather than liquid feeding treatment (P = 0.04). Dry feeding treatment had tendency to increase litter weight at 21d of lactation (P = 0.06) and litter weight gain (P = 0.04) during lactation period (0-3 week). Sows fed dry feeding method made milk containing high content of casein and total solid rather than sows fed liquid feeding method (P = 0.04). In addition, dry feeding treatment had tendency to higher content of milk fat, protein and solid not fat on 21d of lactation (P = 0.07). Sows fed dry feeding type also showed higher milk energy content in milk of 21d lactation (P = 0.05). Furthermore, liquid feeding treatment showed high occurrence in feed waste during lactation period (P <0.01). Conclusion: Dry feeding method was more suitable feeding method to lactating sows under high temperature environment like lactating barn.
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