• Asian-Australasian Journal of Animal Sciences
• /
• 제21권3호
• /
• 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.

• 한국잠사곤충학회지
• /
• 제34권1호
• /
• pp.41-48
• /
• 1992

해충의 미생물적 방제를 위하여 Bacillus thuringiensis 살충제(BT제) 개발에 관한 기초자료를 얻고자 3균주의 공시균주인 B. thuringiensis var. kurstaki, B. thuringiensis var. dendrolimus, B. thuringiensis var. aizawai와 3종의 공시충으로 누에, 흰불나방, 배추흰나비의 유충을 이용하고 B. thuringiensis 내독소단백질을 alkalidyddor 또는 공시된 3종의 곤충소화액으로 처리하여 전기영동을 비교하여 아래와 같은 결과를 얻었다. 1. 3균주의 B. thuringiensis에서 생산된 내독소단백질은 alkali 또는 곤충의 소화액 처리에 따라서 변화되는 뚜렷한 차이가 없는 것으로 나타났다. 2. 각 균주의 내독소단백질의 분자량은 B. thuringiensis var. aizawai가 130kDa, B. thuringiensis var. kurstaki가 140kDa, 130kDa, 그리고 B. thuringiensis var. dendrolimus가 140kDa의 부분에 2band로 나타났으며, 이들 내독소단백질은 alkali 용액이나 숙주곤충의 소화액으로 처리시 활성화되어 40-65kDa의 저분자 단백질로 전환되었다. 또한 숙주곤충소화액에 따라 내독소단백질의 활성화 속도에 약간의 차이가 있는 것으로 확인되었다.

• 동의생리병리학회지
• /
• 제18권4호
• /
• pp.1207-1212
• /
• 2004

Twenty-one diabetic nephropathy patients with normal serum BUN(Blood Urea Nitrogen), creatinine levels and ten chronic renal failure patients with abnormal high BUN, creatinine levels were investigated to evaluate the renal function change after long term herb medicine administration. The hospitalized patients were administrated three times a day with herb medicine, which were prescribe frequently in practical oriental medicine such as many hospital and local clinics. Blood Urea Nitrogen, creatinine and glomerular filtration rate (GFR) were measured immediately after 7days medication. Serum BUN, creatinine levels in diabetic nephropathy patients changed from 17.63±4.38㎎/㎗, 1.09±0.26㎎/㎗(mean±SD) of pre-medication levels to 14.13±3.24 1,20±0.37, 14.75±2.21 1.23±0.55, 12.34±2.89 1.18±0.42 at 7th, 14th, 21th days after herb medicine administration respectively. Also 24hr urine total protein changed from 632.25±254.43㎎/㎗ of pre-medication levels to 623.18±231.56㎎/㎗ after herb medicine administration(P>0.05). Serum BUN, creatinine levels and GFR in chronic renal failure patients changed from 67.45±13.86㎎/㎗, 6.74±2.91㎎/㎗, 13.73±4.21㎖/min pre-medication levels to 61.23±17.75 6.43±2.29 15.49±3.56, 58.84±19.36 5.83±2.51 16.38±2.85, 56.39±20.33 5.64±2.52 16.73±3.40 at 7th, 14th, 21th days after herb medicine administration respectively. Therefore, there was not clinically remarkable difference in the serum BUN, creatinine, GFR levels between pre-medication and post-medication in both Group.

• 한국초지조사료학회지
• /
• 제15권4호
• /
• pp.238-244
• /
• 1995

The objective of this study is to obtain the basic data for investigating the effects of organic reserves on winter survial or regrowth yield. Forage rape(Brassica napus L.) was sown on Sep. 1, 1994. Experimental plots were divided into three replicates under 6 different cutting dates(l0 days interval from Oct. 15 to Dec. 4). Field-grown palnts were sampled on the each cutting date and on the wintering period (Jan. 16) to analyze the nitrogen and non-structural cahohydrate reserves. The rate of winter survival and regrowth yield were also measured in the spring of next year. On the before wintering, dry matter yields were 152, 274, 500, 718, 776 and 981 kg/lOa, respectively, from the cutting date on Oct. 15, Oct. 25, Nov. 4, Nov. 14, Nov. 24, and Dec. 4. Cmde protein yield significantly increased as cutting date was later until Nov. 14, thereafter a significant increase did not occured. Nitrogen and starch contents per plant significnatly increased as the cutting date was later. The increasing rate of starch was greatly higher than that of nitrogen. On the wintering period, nitrogen reserves in mts were 85.3, 68.8, 47.6, 28.3, 44.3, and 55.3 mglplant, and starch reserve were 11 1.3, 75.3, 39.3, 19.6, 26.4 and 34.6 mglplant, respectively, in the plots cut on Oct. 15, Oct. 25, Nov. 4, Nov. 14, Nov. 24, and Dec. 4. It showed that carbohydrate reserves were much highly utilized than nitrogen reserves during wintering period. The rates of winter survival were 91, 83, 46, 22, 35 and 43% and regrowth yields were 692, 545, 316, 84, 127 and 140 kgD.M/lOa, respectively, in each plots. The highly significant correlation (p<0.01) between the level of organic reserves and the rate of winter survival or regrowth yield were obtained.

• Asian-Australasian Journal of Animal Sciences
• /
• 제18권10호
• /
• pp.1375-1378
• /
• 2005

The objective of this study was to investigate polymorphisms of BMPRIB (bone morphogenetic protein type IB receptor) gene and its effect on litter size traits in sheep. Three populations including 101 Small Tailed Han sheep, 79 Poll Dorset and 81 hybrids (Poll Dorset${\times}$Small Tailed Han sheep) were used to detect the polymorphisms in exon 6 region of sheep BMPRIB gene. A fragment of approximately 190bp was amplified by one pair of primers, the polymorphism was revealed from the analysis of three populations by the technique of PCR -SSCP, and a mutation from A to G at 746 of the coding region was confirmed by sequencing in several individual. Statistical results indicated the distribution of allele B (with a A$\longrightarrow$G mutation) and A (without mutation) or genotype AA, AB and BB frequencies differed in three populations. BB genotype (44.55%) and B allele (66.34%) frequencies of Small Tailed Han sheep were higher than those of the others. Analysis of variance showed that the polymorphism of BMPRIB gene was associated with positive effect on litter size traits. The means of genotype BB and AB were about 1.04 and 0.74 more than genotype AA for litter size (p<0.05). Analysis of BMPRIB genotype effects on litter size in three populations indicates the existence of genotype BB or B allele increases the litter size. It suggested that the polymorphism in exon 6 (at 746 in the coding region) of sheep BMPRIB gene may be used as a marker for early selection of prolificacy in sheep.

• 한국자기공명학회논문지
• /
• 제16권1호
• /
• pp.34-45
• /
• 2012

Melanocortin-4 receptor (MC4R) subtype is associated with obese humans. Especially, in a patient with severe early-onset obesity, novel heterozygous mutation in the MC4R gene was detected, resulting in an exchange of aspartic acid to asparagine in $90^{th}$ amino acid residue located in the predicted second trans-membrane domain (TM2). Mutations in the melanocortin-4 receptor (MC4R) gene are the most frequent monogenic causes of severe obesity which have been described as heterozygous with loss of function. In order to compare structure difference between MC4R wild type (MC4R-TM2-wt) and mutant (MC4R-TM2-D90N), we designed both MC4R-TM2-wt and MC4R-TM2-D90N construct in pET 21b vector. In this study, we optimized high-yield purification procedure for recombinant TM2-D90N. Eluted recombinant protein was resolubilized under urea condition for thrombin cleavage reaction and we conducted the high-performance liquid chromatography (HPLC) with reverse phase column under 1% acetonitrile, 0.01% TFA buffer solution. The molecular size of purified target peptide was confirmed by Tricine-SDS page analysis. To characterize MC4R-TM2-D90N, we have performed $^{15}N$-isotope labeling of peptide using M9 media and purified labeled target peptide for hetero-nuclear NMR spectroscopy.

• IMMUNE NETWORK
• /
• 제2권4호
• /
• pp.248-255
• /
• 2002

Background: TAP1 and TAP2 are two ABC transporter genes located within the class II region of the human MHC. Their protein products form a heterodimer whose function is to transport peptides from the cytoplasm into the endoplasmic reticulum. This study was performed to examine the polymorphism of TAP genes and the distribution of HLA-TAP haplotypes in the Korean population through family analysis. Methods: The subjects used in this study were 50 healthy Korean families consisting of 233 individuals. TAP1 (codons 333 and 637) and TAP2 (codons 379, 565, 577, 651, 665, and 687) typings were carried out by the PCR-restriction fragment length polymorphism (RFLP) method. HLA-DRB1 and DQB1 genotyping results from a previous study were used for HLA-TAP haplotype analysis. Results: The number (gene frequency) of TAP1 and TAP2 alleles detected were 3 for TAP1 (A 81.5%, B 17.0%, and C 1.5%) and 8 for TAP2 (A1 32.0%, A2 12.5%, B 34.0%, Bky2 6.5%, C 7.0%, D 3.0%, E 4.5%, and G 0.5%). Eleven TAP1-TAP2 haplotypes were observed with $frequency{\geq}1%$, among which 4 haplotypes (A-B, B-A1, A-Bky2, and C-E) showed weak but significant positive linkage disequilibrium (P<0.05). When DRB1-DQB1 haplotypes were extended to TAP1 and TAP2 loci, much diversification of haplotypes was observed: 19 different DRB1-DQB1 haplotypes formed 58 different haplotypes extended to TAP1 and TAP2 loci. These results add more evidence to the view that recombination hotspot is present within and around TAP gene region. Conclusion: The allele frequencies of TAP1 and TAP2 genes and the distribution of TAP1-TAP2 and HLA-TAP haplotypes were studied in Koreans based on a family study.

• BMB Reports
• /
• 제41권1호
• /
• pp.29-34
• /
• 2008

The aim of this study was to determine whether single nucleotide polymorphisms (SNP) in the beef cattle adipocyte fatty-acid binding protein 3 and 4 (FABP3 and FABP4) genes are associated with carcass weight (CW) and back fat thickness (BF) of beef cattle. By direct DNA sequencing in 24 unrelated Korean native cattle, we identified 20 SNPs in FABP3 and FABP4. Among them, 10 polymorphic sites were selected for genotyping in our beef cattle. We performed SNP, haplotype and linkage disequilibrium studies on 419 Korean native cattle with the 10 SNPs in the FABP genes. Statistical analysis revealed that 220A>G (I74V) and 348+303T>C polymorphisms in FABP4 showed putative associations with BF traits (P=0.02 and 0.01, respectively). Our findings suggest that the polymorphisms in FABP4 may play a role in determining one of the important genetic factors that influence BF in beef cattle.

• 한국축산식품학회지
• /
• 제40권3호
• /
• pp.362-376
• /
• 2020

Rabbit meat bekasam is a traditional fermentation product from Indonesia. This study aimed to determine the chemical and microbiological characteristics of rabbit meat bekasam during the fermentation process in order to isolate, characterize (in vitro and in vivo), and identify lactic acid bacteria (LAB) as the probiotic candidate. The chemical contents of bekasam on 7-day fermentation were investigated in explorative and experimental methods in a completely randomized design. A proximate analysis reported a decrease in the moisture content, fat and carbohydrate content, and an increase in protein content. Also, lactic acid content was increased from 0.48% to 1.12%, and pH was decreased from 5.3 to 4.3. Other properties indicated different values, such as bacteria (2.75×10⁶ to 4.45×10⁷ CFU/g), total LAB (3.82×10⁶ to 4.67×10⁸ CFU/g), total yeast (9.89×10⁶ to 3.82×10⁸ CFU/g) and total mould (4.34×10¹ to 4.86×10³ CFU/g). The experiment produced nine LAB isolates, including two probiotics subjected to further 16S rRNA gene analysis, which indicated that Lactobacillus buchneri was the potential probiotic isolate. After being tested on BALB/c mice, L. buchneri could improve the immune system by inhibiting the growth of Coliform and Salmonella.

• Asian-Australasian Journal of Animal Sciences
• /
• 제13권5호
• /
• pp.686-693
• /
• 2000

Biomass yield, nutritive value and nutrient utilization efficiency of different genera of duckweed (DW, Lemnaceae) under the same nutritional and management conditions were studied. Three genera of DW namely: Lemna perpusilla, Spirodela polyrhiza and Woljfia arrhiza, were cultivated in 18 plastic sinks each of $59{\times}54{\times}35cm^3$ size. Each sink contained approximately 80 L of water and was charged once with 6.648 kg of anaerobically fermented cow dung effluent. The seed rate for Spirodela, Lemna and Wolffia were 600, 400 and $600g/m^2$ respectively. Duckweed were harvested at 48 hours intervals. Media total N concentration for Lemna and Wolffia gradually increased with time. Growth of duckweed was measured by subtracting the inoculum from the total biomass production. Growth of Spirodela ceased within 4 days but Lemna and Wolffia continue to grow up to 34 days. Bio-mass yield was significantly (p<0.05) higher in Wolffia (906 kg/ha/d) than Lemna (631 kg/ha/d). Dry matter (DM) yield was very similar in Lemna (14.80 kg/ha/d) and Wolffia (14.57 kg/ha/d). The N content was non-significant higher in Lemna (5.45%) than Wolffia (5.00%) and Spirodela (4.6%). The crude protein (CP, $N{\times}6.25$) yield was non-significantly higher in Lemna (4.83 kg/ha/d) than Wolffia (4.32 kg/ha/d). The acid detergent fibre (ADF) content was the highest in Wolffia (28.59%), followed by Spirodela (19.47%) and Lemna (12.39%). Utilization efficiency of CP was 273 and 314% respectively for Wolffia and Lemna. However, similar efficiency values for Spirodela was only 1.5%. Considering the bio-mass yield, nutritive value and nutrient utilization efficiency, production performance of DW were in the order of Lemna>Wolffia>Spirodela under the present experimental conditions.