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Studies on the Cryopreservation and Survival after Thawing of Hamster Embryos (Hamster 수정란(受精卵)의 동결보존(凍結保存) 및 생존성(生存性)에 관한 연구(硏究))

  • Oh, Won Jin;Lee, Kyu Seung;Kim, Young Mook
    • Korean Journal of Agricultural Science
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    • v.13 no.1
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    • pp.82-89
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    • 1986
  • This experiment was carried out to determine the optimum freezing and thawing rates of the hamster embryos. The female hamsters were induced to superovulate by intraperitoneal injections of 30 i.u. PMSG and mated with males of the same strain of 4 days the PMSG injection. They were killed and embryos were flushed from the oviduct and uterine horn on 3 days after mating. Embryos were flushed with a modified Dulbecco's phosphate-buffered saline and equilibrated with 1.5 M-dimethylsulphoxide by a 3-step procedure. The freezing rates of the samples were $1^{\circ}C/min$ from room temperature to $-6^{\circ}C$ and the samples were seeded at $-6^{\circ}C$. After being held for 3 min at the seeding temperature, the rates were $0.3^{\circ}C/min$ from $-6^{\circ}C$ to $-35^{\circ}C$. From $-35^{\circ}C$ to $-70^{\circ}C$, the rates were divided into $0.1^{\circ}C/min$, $1^{\circ}C/min$ and $10^{\circ}C/min$, respectively. At $-70^{\circ}C$ the samples were plunged directly into liquid nitrogen. The samples were thawed at $4^{\circ}C/min$ and $12^{\circ}C/min$ from $-196^{\circ}C$ to $37^{\circ}C$, and for 2 min in $37^{\circ}C$ water bath, respectively. The average numbers of ovulation points and embryos recovered were 35.1 and 27.0 appearing 77.0% recovery rates. Eight cell embryos in the embryos recovered were 24.8. The survival rates of embryos according to the freezing rates were 55.5~67.7% at $0.1^{\circ}C/min$, 58.8~64.9% at $1^{\circ}C/min$ and 40.5~44.7% at $10^{\circ}C/min$, respectively. The survival rates at $10^{\circ}C/min$ were significantly low. The survival rates of embryos according to the thawing rates were 53.5% at $4^{\circ}C/min$, 53.7% at $12^{\circ}C/min$ and 59.1% in $37^{\circ}C$ water bath. The survival rates, in $37^{\circ}C$ water bath were slightly higher, but we did not find any differences among them. In conclusion, the best freezing rates of hamster embryos were $1^{\circ}C/min$ from the room temperature to $-6^{\circ}C/min$, $0.3^{\circ}C/min$ from $-6^{\circ}C/min$ to $-35^{\circ}C$ and $-0.1^{\circ}C/min$ or $1^{\circ}C/min$ from $-35^{\circ}C$ to $-70^{\circ}C$. The hamster embryos thawed for 2 min in $37^{\circ}C$ water bath showed the best survival rates.

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Factors Affecting Pregnancy Rates on Transfer of Pronuclear Microinjected Embryos in Korean Black Goats (전핵 미세 주입법으로 생산된 한국흑염소 수정란의 이식 조건이 수태율에 미치는 영향)

  • Choi, Y.S.;Shin, H.G.;Jang, S.K.;Yang, H.S.;Lee, O.K.;Lee, D.S.;Cho, J.K.;Shin, S.T.
    • Journal of Embryo Transfer
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    • v.22 no.1
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    • pp.53-61
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    • 2007
  • This study was investigated factors affecting the pregnancy rates after transfer of pronuclear microinjected embryos for the production of transgenic Korean black goats. Embryo transfer was carried out in 343 recipient Korean black goats from September 1999 to June 2000. Estrus was induced by the insertion of intravaginal progesterone devices $CIDR^(R)$ for 2 weeks. A single injection of 400 IU equine chorionic gonadotropin was administered at 48h before $CIDR^(R)$ removal to increase the proportion of does cycling and ovulation rate. Good quality embryos were prepared by microinjection of DNA into the pronuclei of fertilized goat oocyte and cultured in vitro. Pronuclear microinjected $1{\sim}8$ cell stage embryos were surgically transferred into the oviducts of the recipient at day 4 or 5 following $CIDR^(R)$ removal, and morula to blastocyst stage embryos were surgically transferred into uterus at day 9. Pregnancy was diagnosed by transrectal ultrasound scanning at $20{\sim}30d$ and 8 weeks following embryo transfer. The pregnancy rate was affected by several factors, such as estrus induction, the number of previous transfer, transfer site, stage of CL (corpus luteum), the number of recipient CL, stage of embryos and the number of transferred embryo. The pregnancy rate was significantly higher in recipients that came into estrus naturally than recipients that induced to come into estrus with $CIDR^(R)$(59.1% vs. 36.8%; P<0.05). The pregnancy rate was higher when the embryos were transferred into the left oviduct than transferred into the right oviduct (42.9% vs. 35.3%; P<0.05). The pregnancy rate of recipients with $CH_1$ (early) stage corpus hemorrhagicum in ovary was hi틴or than recipient with $CH_3$ (late) stage hemorrhagicum (47.5% vs. 17.9%; P<0.01). Higher pregnancy rates were obtained by transfer of 1-cell stage embryos into oviduct while late blastocysts (51.6% vs. 66.7%; P<0.01) into uterus. The pregnancy rates when 3 embryos were transferred to recipients were significantly higher than when 2 embryos we.e transferred (47.6% vs. 27.0%; P<0.05). Although there were no significant difference among the group, adhesion of reproductive organs, uterine size, ovulation rate of recipients, presence of large follicle and difficulty of transfer affected pregnancy rate of recipient. Higher pregnancy rates were obtained in the recipients with $8{\sim}15m$ diameter uterine horn as compared to the recipients with <5m diameter or >20mm diameter uterine hem (38.9%, 20% vs. 18.2%), in the recipients with large follicle in the ovulated ovary ipsilaterally (53.6% vs. 37.1%) and in the transfer which was carried out easily (39.2% vs. 27.8%, 0%). In conclusion, the high rate of pregnancy was achieved following transfer of pronuclear microinjected embryos when three or four 1-cell stage embryos were transferred into oviduct with $CH_1$ stage corpus hemorrhagicum in the ovary of recipient which came into estrus naturally.

Studies of the Effects of Acupuncture Stimulation at Huatuo Jiaji(EX B2) Points on Axonal Regeneration of Injured Sciatic Nerve in the Rats (화타협척혈 침자극에 의한 손상 말초신경의 재생효과에 관한 연구)

  • Kim, Dae-Feel;Park, Young-Hoi;Keum, Dong-Ho
    • Journal of Korean Medicine Rehabilitation
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    • v.18 no.4
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    • pp.39-61
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    • 2008
  • Objectives : The present study was performed to investigate whether acupuncture stimulation in the rats affected regeneration properties of the injured sciatic nerve. A differential effect of acupuncture stimulation on the one point near the spinal nerve root controlling sciatic nerve activity and the other point in the peripheral area subordinated by injured nerve was compared. Materials and Methods: Rat sciatic nerves were injured by crush, and the effects on axonal regeneration on injured sciatic nerves were evaluated by acupuncture stimulation at two different regions. In proximal acupuncture stimulation group, acupuncture stimulation was performed on Huatuo Jiaji(EX B2) points located from L5 to S1 vertebral levels to stimulate the nearest spinal nerve root that innervates sciatic nerves. In distal acupuncture stimulation group, acupuncture stimulation was performed on Zusanli(ST 36) and Weizhong(BL 40) points to stimulate at peripheral area dominated by injured sciatic nerves. Acupuncture stimulation was given every other days for 1 or 2 weeks. Sciatic nerve tissues collected from acupuncture stimulation experimental groups, injury control group, and intact animal group were used for protein analysis by Western blotting or Hoechst nuclear staining. To determine axonal regeneration, Dil fluorescence dye was injected into the sciatic nerve 0.5 cm distal to the injury site in individual animal groups and Dil-labeled cells by retrograde tracing were measured in the DRG at lumbar 5 or in the spinal cord. DRG sensory neurons prepared from individual animal groups were used to measure the extent of neurite outgrowth and for immunofluorescence staining with anti-GAP-43 antibody. Results : Animal groups given proximal or distal acupuncture stimulation showed upregulation of GAP-43 and Cdc2 protein levels in the sciatic nerve at 7 days after injury. Cdk2 protein levels were strongly induced by nerve injury, but did not show changes by acupuncture stimulation. Phospho-Erk1/2 protein levels were elevated by acupuncture stimulation above those present in the injury control animals. These increase in regeneration-associated protein levels appeared to be related with increase cell proliferation in the injured sciatic nerves. Hoechst 33258 staining of sciatic nerve tissue to visualize nuclei of individual cells showed increased Schwann cell number in the distal portion of the injured nerve 7 and 14 days after injury and further increases by acupuncture stimulation particularly at the proximal position. Measurement of axonal regeneration by retrograde tracing showed significantly increased Dil-labeled cells in proximal acupuncture stimulation group compared to distal acupuncture stimulation group and injury control group. Finally, an evaluation of axonal regeneration by retrograde tracing showed increased number of Dil labeled cells in the DRG at lumbar 5 or in the ventral horn of the spinal cord at lower thoracic level at 7 days after nerve injury. Conclusions : The present data show that the proximal acupuncture stimulation at Huatuo Jiaji(EX B2) points governing injured sciatic nerves was more effective for axonal regeneration than the distal acupuncture stimulation. Further studies on functional recovery or associated molecular mechanisms should be critical for developing animal models and clinical applications.

The recent essay of Bijeung - Study of III- (비증(痺證)에 대(對)한 최근(最近)의 제가학설(諸家學說) 연구(硏究) - 《비증전집(痺證專輯)》 에 대(對)한 연구(硏究) III -)

  • Yang, Tae-Hoon;Oh, Min-Suk
    • Journal of Haehwa Medicine
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    • v.9 no.1
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    • pp.513-545
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    • 2000
  • I. Introduction Bi(痺) means blocking. It can reach at the joints or muscles or whole body and make pains. Numbness and movement disorders. BiJeung can be devided into SilBi and HeoBi. In SilBi there are PungHanSeupBi, YeolBi and WanBi. In HeoBi, there are GiHyeolHeoBi, EumHeoBi and YangHeoBi. The common principle for the treatment of BiJeung is devision of the chronic stage and the acute stage. In the acute stage, BiJeung is usually cured easily but in the chronic stage, it is difficult. In the terminal stage, BiJeung can reach at the internal organs. BiJeung is one kind of symptoms making muscles, bones and jonts feel pain, numbness or edema. For example it can be gout or SLE etc. Many famous doctors studied medical science by their fathers or teachers. So the history of medical science is long. So I studied ${\ll}Bijeungjujip{\gg}$. II. Final Decision 1. BanSuMun(斑秀文) thought that BiJeung can be cured by blocking of blood stream. So he insisted that the important thing to cure BiJeung is to improve the blood stream. He usually used DangGuiSaYeokTang(當歸四逆湯), DangGuiJakYakSanHapORyeongSan, DoHong-SaMulTang(桃紅四物湯), SaMyoSanHapHeuiDongTang and HwangGiGyeJiOMulTang. 2. JangGeonBu(張健夫) focused on soothing muscles and improving blood seam. So he used many herbs like WiRyeongSeon(威靈仙), GangHwal(羌活), DokHwal(獨活), WooSeul(牛膝), etc. Especially he pasted wastes of the boiled herbs. 3. OSeongNong(吳聖農) introduced four rules to treat arthritis. So he usually used SeoGak-SanGaGam(犀角散加減), BoYanHwanOTang(補陽還五湯), ODuTang(烏頭湯), HwangGiGyeJiOMulTang. 4. GongJiSin thought disk hernia as one kind of BiJeung. And he said that Pung can hurt upper limbs and Seup can hurt lower limbs. He used to use GyeJiJakYakJiMoTang(桂枝芍藥知母湯). 5. LoJiJeong(路志正) introduced four principles to treat BiJeung. He used BangPungTang(防風湯), DaeJinGuTang) for PungBi(風痺), OPaeTang(烏貝湯) for HanBi(寒痺), YukGunJaTang(六君子湯) for SeupBi(濕痺) and SaMyoTang(四妙湯), SeonBiTang(宣痺湯), BaekHoGaGyeTang(白虎加桂湯) for YeolBi(熱痺). 6. GangChunHwa(姜春華) discussed herbs. He said SaengJiHwang(生地黃) is effective for PungSeupBi and WiRyungSun(威靈仙) is effective for the joints pain. He usually used SipJeonDaeBoTang(十全大補湯), DangGuiDaeBoTang(當歸大補湯), YoukGunJaTang(六君子湯) and YukMiJiHwanTang(六味地黃湯). 7. DongGeonHwa(董建華) said that the most important thing to treat BiJeung is how to use herbs. He usually used CheonO(川烏), MaHwang(麻黃) for HanBi, SeoGak(犀角) for YeolBi, BiHae) or JamSa(蠶沙) for SeupBi, SukJiHwang(熟地黃) or Vertebrae of Pigs for improving the function of kidney and liver, deer horn or DuChung(杜沖) for improving strength of body and HwangGi(黃?) or OGaPi(五加皮) for improving the function of heart. 8. YiSuSan(李壽山) devided BiJeung into two types(PungHanSeupBi, PungYeolSeupBi). And he used GyeJiJakYakJiMoTang(桂枝芍藥知母湯) for the treatment of gout. And he liked to use HwanGiGyeJiOMulTangHapSinGiHwan 枝五物湯合腎氣丸) for the treat ment of WanBi(頑痺). 9. AnDukHyeong(顔德馨) made YongMaJeongTongDan(龍馬定痛丹)-(MaJeonJa(馬錢子) 30g, JiJaChung 3g, JiRyong(地龍) 3g, JeonGal(全蝎) 3g, JuSa(朱砂) 0.3g) 10. JangBaekYou(張伯臾) devided BiJeung into YeolBi and HanBi. And he focused on improving blood stream. 11. JinMuO(陳茂梧) introduced anti-wind and dampness prescription(HoJangGeun(虎杖根) 15g, CheonChoGeun 15g, SangGiSaeng(桑寄生) 15g, JamSa(蠶絲) 15g, JeMaJeonJa(制馬錢子) 3g). 12. YiChongBo(李總甫) explained basic prescriptions to treat BiJeung. He used SinJeongChuBiEum(新定推痺陰) for HaengBi(行痺), SinJeongHwaBiSan(新定化痺散) for TongBi(痛痺), SinJeongGaeBiTang(新定開痺湯) for ChakBi(着痺), SinJeongCheongBiEum(新定淸痺飮) for SeupYeolBi(濕熱痺), SinRyeokTang(腎瀝湯) for PoBi(胞痺), ORyeongSan for BuBi(腑痺), OBiTang(五痺湯) for JangBi(臟痺), SinChakTang(腎着湯) for SingChakByeong(腎着病). 13. HwangJeonGeuk(黃傳克) used SaMu1SaDeungHapJe(四物四藤合制) for the treatment of a acute arthritis, PalJinHpPalDeungTang(八珍合八藤湯) or BuGyeJiHwangTangHapTaDeungTang(附桂地黃湯合四藤湯) for the chronic stage and ByeolGapJeungAekTongRakEum(鱉甲增液通絡飮) for EumHeo(陰虛) 14. GaYeo(柯與參) used HwalRakJiTongTang(活絡止痛湯) for shoulder ache, SoJongJinTongHwalRakTank(消腫鎭痛活絡湯) for YeolBi(熱痺), LiGwanJeolTang(利關節湯) for ChakBi(着痺), SinBiTang(腎痺湯) for SinBi(腎痺) and SamGyoBoSinHwan(三膠補腎丸) for back ache. 15. JangGilJin(蔣길塵) liked to use hot-character herbs and insects. And he used SeoGeunLipAnTang(舒筋立安湯) as basic prescription. 16. RyuJangGeol(留章杰) used GuMiGangHwalTang(九味羌活湯) and BangPungTang(防風湯) at the acute stage, ODuTang(烏頭湯) or GyeJiJakYakJiMoTang(桂枝芍藥知母湯) for HanBi of internal organs, YangHwaHaeEungTang(陽和解凝湯) for HanBi, DokHwalGiSaengTang(獨活寄生湯), EuiYiInTang(薏苡仁湯) for SeupBi, YukGunJaTang(六君子湯) for GiHeoBi(氣虛痺) and SeongYouTang(聖兪湯) for HyeolHeoBi(血虛痺). 17. YangYuHak(楊有鶴) liked to use SoGyeongHwalHyelTang(疏經活血湯) and he would rather use DoIn(桃仁), HongHwa(紅花), DangGui(當歸), CheonGung(川芎) than insects. 18. SaHongDo(史鴻濤) made RyuPungSeupTang(類風濕湯)-((HwangGi 200g, JinGu 20g, BangGi(防己) 15g, HongHwa(紅花) 15g, DoIn(桃仁) 15g, CheongPungDeung(靑風藤) 20g, JiRyong(地龍) 15g, GyeJi(桂枝) 15g, WoSeul(牛膝) 15g, CheonSanGap(穿山甲) 15g, BaekJi(白芷) 15g, BaekSeonPi(白鮮皮) 15g, GamCho(甘草) 15g).

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Studies on Direct Sowing-Dry Paddy Rice Culture in the Middle Part of Korea (중부지방에 있어서의 수도건답직파재배 기술체계확립에 관한 시험연구)

  • Jai-Hyoun Lee
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.7 no.1
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    • pp.1-29
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    • 1969
  • Experiments on rice concerning it s varieties, fertilization, seedling dates and herbicides have been conducted to determine the most desirable method of direct sowing cultivation on dry paddy field land in the middle part of Korea. The results obtained at the Office of Rural Development of Choongnam Province are as follows:. 1. Sixteen different derivatives from the main varieties of low land rice were cultivated on a dry paddy field by the direct sowing method; at the same time, a few varieties were tried adopting the common transplanting cultivation method. The yield and yield factors from these two groups were examined to give the following results: a) Compared to the common transplanting cultivation, the direct sowing method showed remarkably increased number of panicles while the number of flowers per panicle was shown to be significantly decreased. The maturing ratio was detected to be lowered. The yield horn them differed according to the different varieties : good yield was obtained from Hokwang while Norin #25 proved poor when compared with the common transplanting cultivation method. b) Among sixteen varieties tested, Sunsou, Norin #25, Jaigou, Hokwang, Palkueng and Gosi showed comparatively high yields, their yield being more than 325 kilograms per 10 Are, but Nampoong, Paldal, Nongkwang, Norin #29, Eunbangju #101 and Shiro gane showed less yield, their yield being less than 271 kilograms per 10 Are, the relations between the yield and the yield factors can be summarized as follows; Number of varieties and their rice yield. 1) The varieties which were great in the, number of panicles and high in yield=Jaigoun, Hokwang Palkueng and Gosi. 2) The varieties which were low in the number of panicles and high in yield=Sounsou and Norin #25. 3) The varieties which were great in the number of panicles and poor in yield=Eunbangju #101 and Sirogane. 4) The varieties which were poor in the number of panicles and poor in yield: Nampung, Paldal and Norin #29. Number of flowers per panicle and yield. 1) The varieties which were great in the number of flowers per panicle and high in yield: Sounsou, Norin #25 and Gosi. 2) The varieties which were poor in the number of flowers per panicle and high in yield ; Jaigoun, Hokwang and Palkueng. 3) The varieties which were great in the number of flowers per panicle and poor in yield: Paldal and Nampung. 4) The varieties which were poor in the number of flowers per panicle and poor in yield: Norin #29. Eunbangju #101 and Sirogane. Maturing ratio and yield. 1) The varieties which were high in the maturing ratio and high in yield: Jaigoun, Sounsou, Norin #25 and Palkueng. 2) The varieties which were low in the maturing ratio and high in yield: Hokwang and Gosi. 3) The varieties which were early maturing rat io and low in yield: Hokwang and Gosi. 4) The varieties which were late maturing ratio and poor in yield: Eunbangju #101, Nampungand Sirogane 1, 000 grain weight and yield. 1) The varieties which were heavy in 1, 000 grains weight and high in yield=Norin #25 and Hokwang. 2) The varieties which were light in 1, 000 grains weight and high in yield=Sounsou and Jaigoun. 3) The varieties which were heavy in 1, 000 grains weight and poor in yield=Nongkwang and Eunbanju. 4) The varieties which were light in 1, 000 grains weight and poor in yield=Norin #29 and Sirogane. 2. The experiment on fertilization showed that the most desirable amount to be given per 10 Are was 10 kilograms of Nitrogen, 5 kilograms of phosphate and 6 kilograms of potassium; and when the Nitrogen given exceeded 8 kilograms, its effect was better when given in amsll consecutive (split) amounts, while the maturing ratio and the number of the flowers per panicle increased when Nitrogen was given in large amount during the later stage of growth of rice. 3. The experiment on the date and amount of seedling showed that the tested variety, Sunsou gave the best results when planted on the days between 25 April and 10 May. Eight liters per 10 Are were preferable if planted early and 12 liters per 10 Are if planted late. The reason why the later planting gave a lower yield was that the number of flowers per panicle was fewer. 4. The experiment on the irrigation for rice with direct sowing cultivation immersed in water showed that it was the most satisfactory when irrigated on 25th June, 55 days after its seedling, its plot giving the best yield. The plots 10th June and 15th July showed just as good results. However, irrigated later, than 15th July it showed lower yields. 5. Compared to the yield of the plot controlled by the common method, the yield from the plots treated with chemical herbicide such as LOROX, TOK, PCP, SWEP, Mo-338 on dry condition soil seemed poorer, but significant difference was not found statistically. On the other hand in the case where chemical herbicides such as TOK, Mo-338, Stam F-34 or ORDRAM were used after irrigation, the yield from the ORDRAM and TOK treated plots did not show significant differences compared to the common hand weed controling method, but those treated with chemicals other than the above showed a lower yield.

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