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유리화 동결법에 의한 난자의 동결 중 항동해제에 Cytoskeleton Stabilizer, Taxol의 처리 후 배발달율의 개선에 관한 연구

  • 정형민;박성은;임정묵;고정재;정미경;차광렬
    • Proceedings of the KSAR Conference
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    • 2001.03a
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    • pp.74-74
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    • 2001
  • 난자 동결방법의 선별은 보다 효과적인 난자은행의 개발에 필수 불가결한 중요한 요소이다. 이전의 연구에서 마우스의 난자를 ethylene glycol과 electron microscope grid를 이용한 유리화 동결법으로 동결 융해한 결과 기존의 slow freezing 방법에서보다 높은 생존율과 배발달율이 나타남을 관찰하였다. 그러나 동결융해후의 난자는 방추사와 염색체의 이상성이 대조군에 비해 높은 빈도로 나타나 융해후의 배발달율을 감소시키는 것으로 보고되었다. 이에 본 연구에서는 유리화동결법동안 항동해제에 Cytoskeleton system을 안정화시키는 cytoskeleton stabilizer인 taxol을 첨가시킨후 동결시켰을때 생존율과 발달율을 개선시킬 수 있는지 알아보고자 본 실험을 시행하였다. ICR mouse의 성숙란을 채취하여 연구목적에 따라 taxol을 첨가시키지 않은 대조군과 첨가시킨 실험군으로 분류하였다. 동결방법은 난자를 1.5 M ethylene glycol (EG)에 2분 30초간 노출시킨후 5.5 M EG와 1 M sucrose가 첨가된 동결액에 20초간 노출시킨 후 Grid에 난자를 부착시킨후 직접 액체질소에 침지하여 동결하였다. 동결후 난자는 5단계로 융해를 실시한 후 정자와 체외수정을 시킨 후 수정된 난자는 modified P1 배약액에 124 h까지 발달율을 관찰하였고, 배양 후 발달된 배반포는 대조군과 실험군, 각각 4마리의 발정동기화된 recipient에 이식을 시행하였다. 배발달율은 대조군에 비해 실험군에서 4세포기 (48 vs. 84.4%), 8세포기 (34% vs. 70.6%), 상실배 (26% vs. 58.6%) 그리고 배반포 발달율은 (24% vs. 58.6%)로 높게 관찰되었다. 배아이식후 대조군과 실험군에서 각각 2 마리가 임신이되어 정상적인 산자를 분만하였다. 따라서 항동해제에 taxol의 첨가는 동결 융해후의 난자의 배발달율을 증진시킬 수 있었다..8%로 나타나 난할율 및 배반포 발생율에 있어서 융합조건에 따라 큰 차이는 없었으나 1.9㎸/cm, 30$\mu\textrm{s}$ 2회의 조건이 다른 조건들에 비하여 유의적으로 낮았다. 따라서, 체세포와 수핵란 세포질간의 융합율과 배반포 발생에 미치는 영향은 전압보다는 시간에 더 크게 받음을 알 수 있었으며, 이와 같은 결과에서 융합시 시간을 오래 주는 것보다 전압을 높이는 것이 수핵난자의 세포질에 상해를 줄이고 이후 배반포 발생에 유리할 것으로 사료되었다.면에서도 더욱 더 활발할 것으로 기대된다. 배란후 72시간째에 초음파진단기를 이용하여 난소의 난포발달을 조사한 결과 , 대조구와 bFF처리구에 비해 AI처리구에서 발달난포가 유의적으로 많은 것을 확인하였다. 이상과 같은 결과로, Anti-inhibin serum은 한우 자체에서 분비하는 Inhibin을 특이하게 억제하여 Inhibin에 의해 억제되는 FSH분비가 촉진됨으로써 난포발달과 estrogen의 농도가 촉진되는 것으로 사료되어 anti-inhibin serum이 한우의 과배란유기 효과가 있는 것으로 사료된다.정량 분석한 결과이다. 시편의 조성은 33.6 at% U, 66.4 at% O의 결과를 얻었다. 산화물 핵연료의 표면 관찰 및 정량 분석 시험시 시편 표면을 전도성 물질로 증착시키지 않고, Silver Paint 에 시편을 접착하는 방법으로도 만족한 시험 결과를 얻을 수 있었다.째, 회복기 중에 일어나는 입자들의 유입은 자기폭풍의 지속시간을 연장시키는 경향을 보이며 큰 자기폭풍일수록 현저했다. 주상에서 관측된 이러한 특성은 서브스톰 확장기 활동이 자기폭풍의 발달과 밀접한 관계가 있음을 시사한다.se that were all low in two aspects, named "the Nonsignificant group". And the issues were high risk perception in general setting and

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Recent Advances in Amino Acid and Energy Nutrition of Prolific Sows - Review -

  • Boyd, R.D.;Touchette, K.J.;Castro, G.C.;Johnston, M.E.;Lee, K.U.;Han, In K.
    • Asian-Australasian Journal of Animal Sciences
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    • v.13 no.11
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    • pp.1638-1652
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    • 2000
  • Prolific females require better nutrition and feeding practice because of larger litter size and the substantial decline in body fat. Life-time pig output will be compromised if body protein and fat are not properly managed. First litter females are especially vulnerable because they can loose ${\geq}15%$ of whole-body protein. Conservation of body protein mass during first lactation minimizes wean to estrus interval and increases second litter size (up to 1.2 pigs). The ability to influence litter-size by amino acid nutrition is a new dimension in our understanding. A P2 fat depth below 12 mm at farrow and below 10 mm at wean compromised wean to estrus interval (>2 d) and next litter size (0.5 to 1.5 pigs) in sows. It is now clear that a 'modest' excess of feed during the first 72 h of pregnancy decreases embryo viability so that the potential for an increased litter size at birth is not realized. The capacity for milk production by prolific young sows is 25% higher than the standard used previously (NRC, 1988). First litter females averaged 9.82 kg milk/d for a 21 d lactation. Second and third litter counterparts averaged 10.35 kg/d. Milk production was 95% of peak by 10 d of lactation and sows were in greatest negative energy and lysine balance during the first 6 d. Nearly 45% of the total loss in body protein occurred within the first 6 d, but this could reduced to 30-35% by using a more aggressive feeding strategy after parturition. There appear to be 2 phases in lactation for lysine need (d 2-12 vs 12-21). Feeding to the higher level alleviates the second litter size decline. The lysine requirement for lactation can be predicted with accuracy, but we are not able to predict the second limiting amino acid. Mammary uptake of valine relative to lysine and recent work with practical diets suggest that the recent NRC (1998) pattern is realistic and that threonine and valine could be co-limiting for corn-soy diets for prolific sows nursing 10-11 pigs. Empirical studies are needed to refine the ideal pattern so that synthetic lysine can be used with more confidence. Milk fat output for the elite sow is extraordinary and poses an unnecessarily high energetic cost. Methods that reduce mammary fat synthesis will benefit the sow and may enhance piglet growth.

The Effect of Additional Mobilization with Conservative Physical Therapy in Patients with Frozen Shoulder on ROM and Subjective Pain Scale (동결 견 환자에 대한 보존적 물리치료와 관절 가동술의 병행이 주관적 통증지수와 관절가동범위에 미치는 효과)

  • Hyong, In-Hyouk;Ha, Mi-Sook
    • The Journal of the Korea Contents Association
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    • v.9 no.11
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    • pp.271-279
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    • 2009
  • Frozen shoulder is clinically characterized by pain and reduced ROM. The therapeutic goal of frozen shoulder can be reducing pain and increasing ROM in shoulder joint, resulting in improvement of joint movement. Therefore, this study was conducted to know the effect of mobilization in parallel with conservative physical therapy(H/P, TENS, U/S) in patients with frozen shoulder on subjective pain scale and ROM. 26 patients diagnosed with frozen shoulder were included for study. Among them, 13 patients(experiment group) were managed by mobilization as well as conservative physical therapy, another 13 patients(control group) were only treated by conservative physical therapy alone. All the patients were treated three times a week for 4 weeks, and after each treatment subjective pain scale and ROM were measured. In experimental group, pain has decreased and ROM has increased in abduction, external rotation and internal rotation. In control group, pain has also decreased but ROM has not changed in abduction, external rotation and internal rotation. There were no significant difference in subjective pain scale between two groups but some difference in ROM (external rotation, internal rotation). According to the results, we concluded that both conservative physical therapy and joint mobilization technique are more effective for increasing ROM especially external rotation, internal rotation than conservative physical therapy alone.

Nutrition Survey in a Korean Mountainous Farming Area (산간지(山間地) 농촌주민(農村住民)의 영양실태조사(營養實態調査))

  • Ham, Jung-Rea;Kim, Hyong-Soo;Lee, Ki-Yull;Kim, Young-Hoo
    • Journal of Nutrition and Health
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    • v.6 no.3
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    • pp.37-45
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    • 1973
  • The main character of the Korean diet has been found to be low in protein both quantity and quality and high in carbohydrate. The purpose of this survey was to study the amount of salt intake related to the dietary pattern in Korea. The nutrition survey was conducted in a mountainous farming area located in Auhchun-ri, Gaebuk-myon, Changsoo-gun, Chunbuk Province, February 14-19 in 1973 (7 days). The precise weighing method was used in evaluating the kinds of foods and nutrients intake for 24 households during a three day period. The physical examinations were performed by a doctor on 120 persons and a detailed biochemical test on both blood and urine was made on 42 persons over 40 years old. The results obtained are summarized as follows: (1) Average nutrients intake of an adult per day: calorie intake was 2,446 Cal and its components-protein(61.1g) was 10 percent, fat(12.9g) was 5 percent and carbohydrate(521g) contributed 85 percent of the total calories. Other nutrients-calcium (443mg), thiamine(1.09mg), riboflavin (0.90mg), niacin (14.4mg) and vitamin C (63.2mg) were lower than the recommended daily allowance but vitamin A(2,083 I.U.), iron(11mg) and phosphorous(998mg) were slightly higher than that. (2) To evaluate the nutritional deficiences, clinical examinations were conducted. Angular stomatitis was present in 16.7 percent of those examiners. No edema was found. The rate of osteoarthritis, hepatomegaly diseases appeared in 20 percent of the total subjects and the symptoms appeared highest among those Iron 50 to 59 years old. (3) The following chemical components of blood serum were analyzed and found to be within the normal range: glucose, blood urea nitrogen, uric acid, total protein, albumin, globulin, bilirubin, total cholesterol, inorganic phosphate, alkaline phosphatase, sodium, potassium, chloride, calcium and lecithine dehydrase. One case of each of the following were found: hyperglycemia, hypocholesterolemia, renal problem, hypoproteinaemia and diabetes mellitus, and two persons were classified as showing hypoglycemia and hyponaturemia. (4) The sodium content in urine was 199.6 mEq/L, potassium content was 24.6 mEq/L. The sugar, pH and specific gravity in the urine was shown to be normal.

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Effect of Dendritic Cell Based Cancer Vaccine Using Allogeneic Tumor Cell Lysate in Melanoma Pulmonary Metastasis Model (동종 종양 세포 용해액을 이용한 수지상 세포 항암 백신의 흑색종 폐암 전이 모델에서의 효과 연구)

  • Lee, Young-Joon;Kim, Myung-Joo;In, So-Hee;Choi, Ok-Mi;Baek, So-Young;Kwon, Young-Do;Lee, Hyun-Ah
    • IMMUNE NETWORK
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    • v.5 no.3
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    • pp.163-171
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    • 2005
  • Background: To perform the successful dendritic cell-based cancer immunotherapy one of the main issues to be solved is the source of antigen for DC pulsing. Limitations occur by using auto-tumor lysate due to the difficulties obtaining enough tumor tissue(s) quantitatively as well as qualitatively. In this study the possibility of allogeneic tumor cell lysate as a DC pulsing antigen has been tested in mouse melanoma pulmonary me tastasis model. Methods: B16F10 melanoma cells $(1{\timeS}10^5/mouse)$ were inoculated intra venously into the C57BL/6 mouse. Therapeutic DCs were cultured from the bone marrow myeloid lineage cells with GM-CSF and IL-4 (1,000 U/ml each) for 7 days and pulsed with lysate of either autologous B16F10 (B-DC), allogeneic K1735 (C3H/He origin; K-DC) or CloneM3 (DBA2 origin; C-DC) melanoma cells for 18 hrs. Pulsed-DCs $(1{\times}10^6/mouse)_{[CGP1]}$ were injected i.p. twice with one week interval starting from the day 1 after tumor cell inoculation. Results: Without observable toxicity, allogeneic tumor cell lysate pulsed-DC induced the significantly better anti-tumor response (tumor scale: $2.7{\pm}0.3,\;0.7{\pm}0.3\;and\;0.3{\pm}0.2$ for saline, B-DC and C-DC treated group, respectively). Along with increased tumor specific lymphocyte proliferations, induction of IFN-${\gamma}$ secretion against both auto- and allo-tumor cell lysates was observed from the DC treated mice. (w/B16F10-lysate: $44.97{\pm}10.31,\;1787.94{\pm}131.18,\;1257.15{\pm}48.27$, w/CloneM3 lysate: 0, $1591.13{\pm}1.83,\;1460.47{\pm}86.05pg/ml$ for saline, B-DC and C-DC treated group, respectively) Natural killer cell activity was also increased in the mice treated with tumor cell lysate pulsed-DC ($8.9{\pm}_{[CGP2]}0.1,\;11.6{\pm}0.8\;and\;12.6{\pm}0.7%$ specific NK activity for saline, B-DC and C-DC treated group, respectively). Conclusion: Conclusively, promising data were obtained that allogeneic-tumor cell lysate can be used as a tumor antigen for DC-based cancer immunotherapy.

Estimation of Habitat Suitability Index of Fish Species in the Geum River Watershed (금강수계 하천에서의 어류 서식처적합도지수 산정)

  • Kang, Hyeongsik;Im, Dongkyun;Hur, Jun Wook;Kim, Kyu-Ho
    • KSCE Journal of Civil and Environmental Engineering Research
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    • v.31 no.2B
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    • pp.193-203
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    • 2011
  • With the recent growth of environmental and ecological interests, various river restoration and habitat environment creation projects are being carried out. For this, the estimation of the habitat flow discharge is important. In U.S. and Europe nations, The instream flow incremental methodology (IFIM) has been used to estimate the habitat discharge. IFIM is the method that can be applied to evaluate the flow discharge for the suitable habitat. To use the IFIM in river, a habitat suitability index(HSI) for the target organism is needed. However, HSIs for only two species of Zacco platypus and Zacco temminckii were proposed from the field monitoring. Thus, for the estimation of the ecological flow rate for a group of fish, the development of the HSIs for various fish are necessary. In this study, physical data such as water level and flow rate, chemical data such as acidity and dissolved oxygen, and life data such as fish types and population are collected in Keum river watershed. Based on the 2,736 field data, HSIs for the following 6 fish are developed: Zacco platypus, Zacco temminckii, Microphysogobio yaluensis, Coreoleuciscus splendidus, Pungtungia herzi, Pseudogobio esocinus. Through the comparison with HSIs in the literature, the developed HSIs are modified. Also, the limits of Froude number, pH, and DO for 6 fish are proposed. The HSIs developed in this study can be utilized as a essential data for performing river project evaluations.

The Formation of Serrated Grain Boundaries and Its Influence on Boron Segregation and Liquation Behavior (파형 결정립계 생성이 보론 편석 및 액화거동에 미치는 영향)

  • Hong, H.U.;Kim, I.S.;Choi, B.G.;Yoo, Y.S.;Jo, C.Y.
    • Proceedings of the KWS Conference
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    • 2010.05a
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    • pp.73-73
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    • 2010
  • 합금원소가 다량 첨가된 고합금강, 스테인리스강, Ni기 초내열합금 등은 용접시 혹은 후열처리 동안 열영향부 (HAZ: heat-affected-zone)에서 결정립계를 따라서 액화균열이 종종 발생한다. 이러한 액화균열은 급속한 가열시 HAZ의 결정립계가 국부적으로 용융되어 액상필름을 형성하고, 냉각시 수축으로 인한 인장구속응력에 의해 필름을 따라서 균열이 발생하여 생성된다. HAZ 결정립계 액화는 탄화물, 황화물, 인화물, 보론계 화합물 등이 급가열시 기지와의 반응에 의해 표피 액상을 형성하는 조성적 액화 (constitutional liquation)에 의한 액상의 결정립계 침투로 설명되거나, 결정립계 자체의 용융점을 상당량 낮추는 보론(B), 인(P), 황(S)등의 편석에 의한 국부적 입계 용융으로 주로 연관 지어 해석한다. HAZ 액화균열은 고온 입계균열 현상이므로, 결정립계의 특성에 따라 크게 영향을 받으며 결정립계 character 설계에 의해 액화균열 저항성을 개선시킬 수 있음을 유추할 수 있다. 한편, 본 연구자들은 최근 Ni기 초내열합금에 있어 입계 serration 현상을 새롭게 발견하였으며, 이론적 접근법을 통해 serration을 위한 특별한 열처리 방법을 개발하였다. 형성된 파형입계는 결정학적인 관점에서 조밀 {111} 입계면을 갖도록 분해 (dissociation)되어 낮은 계면에너지를 갖게 됨을 확인하였으며, 입계형상 변화뿐만 아니라 탄화물 특성변화까지 유도하여 크리프 수명을 기존대비 약 40% 정도 향상시킴을 확인하였다. 본 연구에서는 이러한 직선형 입계 대비 'special boundary'로 간주되는 파형입계가 도입될 경우, 보론 편석 및 HAZ 액화거동에 미치는 영향을 고찰하고자 하였다. SIMS (secondary ion mass spectrometry)를 이용하여 열처리 직후 결정립계 보론편석 정도를 비교하였다. 파형입계 시편의 경우, 일반직선형 시편에 비해 결정립계에 보론편석 저항성이 우수함을 확인할 수 있었다. 재현 HAZ 열사이클 시험을 통해 미세조직을 정량적으로 분석하였다. 파형입계 시편 및 일반직선형 시편 모두 최고온도 $1060^{\circ}C$이상부터 입계 탄화물이 기지내로 완전 용해되고 입계가 액화되기 시작하였다. 최고온도별로 입계액화비율을 정량적으로 비교한 결과, 파형입계가 직선입계 대비 훨씬 낮음을 확인할 수 있었으며, 때때로 액화된 필름이 입계를 따라 전파되지 않고 부분적으로 단락되어 있음이 관찰되었다. 액화시험 후 투과전자현미경을 이용한 EDS (energy dispersive spectrometry) 분석을 통해 결정립계 액화의 주요원인은 입계 $M_{23}C_6$의 조성적 액화반응 보다는 보론 편석 (원자 및 $M_{23}(CB)_6$)으로 인한 결정립계 국부용융이 더 유력함을 유추할 수 있었다. 따라서 상기 결과로부터 입계구조가 안정되어 계면에너지가 낮은 파형입계가 보론편석에 대한 저항성이 우수하였으며, 이러한 결과는 액화 저항성에 대응되어 영향을 미침을 알 수 있었다. 게다가 파형입계에 액상 필름이 생성되더라도 낮은 계면에너지에 의해 비롯된 상대적으로 낮은 적심성 (wettability)에 의해 필름이 쉽게 전파되지 않음을 'Smith 입계 wetting 이론'을 이용하여 해석할 수 있었다.

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Purification and Characterization of Degradative Enzyme of Dental Plaque from Streptomyces sp. Y9343 (Streptomyces sp. Y9343이 生産하는 齒面細菌膜 分解酵素의 精製와 特性)

  • Kim, Seong-Joo;Han, Hong-Keun;Yoon, Jeong-Weon
    • Microbiology and Biotechnology Letters
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    • v.24 no.1
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    • pp.9-18
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    • 1996
  • Streptococcus mutans has been implicated as primary causative agents of dental caries by insoluble glucan (IG) in human and experimental animals. An attempt was made to search for the ${\alpha}$-1,3 glucanase that degrades IG produced by S. mutans. ${\alpha}$-1,3 glucanase was detected in the culture supernatant of microorganisms, which are isolated from soils on agar medium containing IG as a sole carbon source. This Streptomyces sp. hydrolysed IG produced by immobilized S. mutans and was named as Y9373. This enzyme required ${\alpha}$-1,3 glucan (IG) as an inducer. The optimum conditions for enzyme production were studied. The enzyme was purified by 30~70% $(NH_4)_2SO_4$ precipitation, anion exchange chroma tography on DEAE-cellulose and gel filtration on Sepadex G-75. The purified enzyme has a specific activity of 7840.0 U/mg protein giving 32.1-fold purification and final yield of 0.53%. The molecular weight was estimated to be about 22.5 kDa by SDS-PAGE. The optimum pH and temperature for enzyme reaction were 6.5 and 37$^{\circ}C$, respectively and the enzyme was relatively stable at the temperature below 60$^{\circ}C$. The activity of purified enzyme was enhanced by adding $Co^{2+},\;Mn^{2+}\;and\;Mg^{2+}$ into the medium, whereas inhibited by adding $Hg^{2+},\;Zn^{2+}$ and SDS. The $K_m\;and\;V_{max}$ value of ${\alpha}$-1,3 glucanase for IG were estimated to be 2.50 mM and 0.0431 mM/min, respectively. The thin layer chromatographic analysis of hydrolysates from IG with ${\alpha}$-1,3 glucanase showed that glucose was the main product of reaction. This enzyme activity was about 14 times higher than marketing dextranase as preventive agent against artificial dental caries by S. mutans in TH medium including 5% sucrose after 30 minutes.

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Effects of Gonadotrophins on In Vitro Growth and Maturation of Mouse Preantral Follicles (생쥐 Preantral Follicles의 체외성장 및 성숙에 있어서 Gonadotrophins의 역할)

  • 김동훈;지희준;강희규;한성원;이훈택;정길생;이호준
    • Korean Journal of Animal Reproduction
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    • v.23 no.1
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    • pp.53-61
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    • 1999
  • The present study was designed to investigate the effects of gonadotrophins on in vitro growth and maturation in mouse preantral follicles. Ovaries were removed from 12-day-old ICR mice. Follicles were dissociated enzymetically in Leibovitz L-15 medium containing 1 mg/$m\ell$ collagenase and 0.2 mg/$m\ell$ DNase I. The follicles were cultured on Transwell-COL membrane inserts in six well cluster dishes for 10 days. The culture medium was $\alpha$MEM medium supplemented with 5% fetal bovine serum and FSH or HMG. After 10 days of growth in vitro, follicles were allowed to mature for 18~20 hr in medium supplemented with 1.5 IU/$m\ell$ hCG. The oocytes were then denuded of their cumulus cells and assessed maturation status. Concentrations of oestradiol and progesterone were measured with a radioimmunoassay. Oocyte diameter was determined with an ocular micrometer. The survival and Metaphase II rates of oocytes were significantly higher in FSH treatment groups than in control group (P<0.001), but there were no differences among the groups of treated FSH concentration. The survival and Metaphase II rates of oocytes in HMG treatment group (60.9 and 40.6%) were higher than in FSH treatment group (76.6 and 48.2%) and control group (49.2 and 7.1%). The survival and Metaphase II rates of oocytes on both FSH and LH treatment groups were no differences among the ratios of FSH and LH. Diameter of oocyte was no differences among the treatment groups, but smaller than compared to in vivo grown oocyte. Through the entire culture period, secretions of oestradiol and progesterone were significantly less in control group than in HMG and FSH treatment groups. These results suggest that gonadotrophins playa key role in in vitro culture of mouse preantral follicles. Especially, addition of FSH and LH should be more effective than FSH alone.

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Allelopathic Effect of Sorghum Extract and Residues on Selected Crops and Weeds (수수의 타감작용(他感作用)에 관(關)한 연구(硏究))

  • Kim, S.Y.;De Datta, S.K.;Robles, R.P.;Kim, K.U.;Lee, S.C.;Shin, D.H.
    • Korean Journal of Weed Science
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    • v.14 no.1
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    • pp.34-41
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    • 1994
  • To better understand the allelopathic effect of sorghum(Sorghum vulgare L.), the inhibitory activities of water extracts of the stem, leaf and root, and of residues of the stem to major crops and weeds associated with them were evaluated. The allelopathic activity of sorghum plants was species specific, and depended on source and concentration. Germination, and shoot and root length of all test species were inhibited by the different concentrations of the stem extract. Among the crop species, radish showed the most inhibition, followed by wheat and rice. Maize was the least sensitive species. Of the weed species, Ipomoea triloba was most inhibited, followed by Echinochloa colona and Rottboellia cochinchinensis. The water extracts of leaves, stems, and roots significantly inhibited germination and seedling growth in E. colona and radish. The stem extract gave the greatest inhibitory effect on E. colona while all three extracts produced similar response in radish. In the greenhouse trial, sorghum stem residue placed on the soil surface as mulch significantly inhibited seedling growth in E. colona and radish, but not that in rice.

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