• 생명과학회지
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• 제24권8호
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• pp.915-926
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• 2014

리보솜은 mRNA상의 유전정보를 단백질로 번역하는 세포에 필수적인 거대복합체이다. 이러한 리보솜은 리보 핵산단백질 복합체로, rRNA와 리보솜 단백질로 이루어져있다. 리보솜 조립과정은 리보솜 단백질 이외에도 많은 조립인자들이 각 구성요소의 조립을 도움으로써 이루어진다. 세포 내 리보솜 조립과정에 참여하는 조립인자들로 GTPase, ATPase, 샤페론, RNA helicase, 수식효소 등 다양한 단백질들이 알려졌다. 리보솜 조립과정 중 이러한 조립인자들은 리보솜 단백질 또는 rRNA의 수식에 참여하거나, 리보솜 단백질들과 rRNA의 조립 등을 돕는다. 이러한 리보솜 조립인자들에 관한 유전학적, 구조적, 생화학적 실험결과들이 많이 존재하지만 정확한 리보솜 조립과정과 이러한 조립인자들의 역할에 대해서는 아직 밝혀지지 않았다. 현재까지의 연구결과를 바탕으로 E. coli의 리보솜 조립과정을 돕는 단백질들에 대하여 알아보고자 한다.

• Clinical and Experimental Reproductive Medicine
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• 제16권1호
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• pp.69-79
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• 1989

Steroid hormone profiles during luteal phase of clomiphene citrate(CC)/human menopausal gonadotropin(hMG)/human chorionic gonadotropin(hCG)-stimulated in vitro fertilization (IVF) cycles and of follicle-stimulating hormone(FSH)/hMG/hCG-stimulated IVF cycles were compared. In seventy three cycles stimulated with CC/hMG/hCG regimen, follicles were aspirated during exploratory laparotomy and yielded 7 pregnancies, and in 83 cycles stimulated with FSH/hMG/hCG regimen, follicles were aspirated by laparoscope and made 13 pregnancies. Serum estradiol($E_2$) and progesterone($P_4$) levels were determined on days 2, 5, 7, and 9 after follicle aspiration. The FSH/hMG/hCG regimen was more effective than the CC/hMG/hCG regimen in folliculogenesis, ie, ovarian stimulation, follicular phase $E_2$ peak levels, oocyte maturation, and the number of retrieved oocytes. There was no significant difference between luteal serum $P_4/E_2$ ratio of the two regimens, suggesting that secretory endometrial build-up ability for implantation may not differ each other. Several significant correlations were observed between follicular phase seum $E_2$ peak levels and luteal phase serum $E_2$ and $P_4$ levels in the FSH/hMG/hCG-stimulated cycles but any correlation was not significant in the CC/hMG/hCG-stimulated cycles, suggesting that somewhat more follicles may eventually fall in atresia even after attaining dominant stage in the CC/hMG/hCG-stimulated cycles than the FSH/hMG/hCG-stimulated cycles.

• 한국수정란이식학회지
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• 제29권3호
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• pp.273-281
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• 2014

This study was carried out to establish the system of OPU derived embryo production, management of recipients as well as offspring production. OPU derived embryo production system was carried out of aspiration of immature oocytes 2 times per week, total 24 times for 3 months by an ultrasonographic guided follicular aspiration system and then produced in vitro-produced blastocysts by in vitro maturation, fertilization and culture system. This work was collected total 13,866 oocytes, average $8.2{\pm}4.5$ oocytes per session and 8,170 G1 + G2 grade oocytes, average 4.8 oocytes per session by 1,692 times session of total 71 donors for 4 years from 2010 to 2013. The rate of cleavage and blastocyst developmental competence were obtained 11,825 (85.3%) and 5,032 (36.3%) that was $7.0{\pm}3.8$ cleaved embryos and $3.0{\pm}2.5$ blastocysts per session. OPU derived embryo transfer were taken place in 2, 4, 6 and 7 local governments at 2010, 2011, 2012 and 2013 for 4 years and pregnancy rate were obtained 41.2, 43.9, 46.5 and 49.7% in each years. It means that pregnancy rate was continuously improved according of every year for 4 years. Pregnancy rate was significantly different according to individual local government in which was 62.7% in B, but 24.2% in F at 2012. Paternity identification was carried out total 26 offspring in C local government of 2012 and then confirmed 100% agreement of its analysis. In conclusion, the results obtained the possibility of mass production of elite cow embryos as well as offspring by OPU derived embryo production system, of which could be decreased the required time of genetic improvement.

• Reproductive and Developmental Biology
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• 제36권2호
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• pp.115-120
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• 2012

This study was conducted to examine the optimal concentration and treatment time of antioxidants for inhibition of the ROS generation in bovine somatic cell nuclear transfer (SCNT) embryos. Bovine oocytes were activated parthenogenetically, during which oocytes were treated with various antioxidants to determine the optimal concentrations and kind of antioxidants. Determined antioxidants were applied to oocytes during in vitro maturation (IVM) and/or SCNT procedures. Finally, antioxidant-treated SCNT embryos were compared with in vitro fertilized (IVF) embryos. $H_2O_2$ levels were analyzed in embryos at 20 h of activation, fusion or insemination by staining of embryos in $10{\mu}M$ 2'7'-dichlorodihydrofluorescein diacetate (H2DCFDA) dye, followed by fluorescence microscopy. $H_2O_2$ levels of parthenogenetic embryos were significantly lower in $25{\mu}M$ ${\beta}$-mercaptoethanol (${\beta}$-ME), $50{\mu}M$ L-ascorbic acid (Vit. C), and $50{\mu}M$ L-glutathione (GSH) treatment groups than each control group ($24.0{\pm}1.5$ vs $39.0{\pm}1.1$, $29.7{\pm}1.0$ vs $37.0{\pm}1.2$, and $32.9{\pm}0.8$ vs $36.3{\pm}0.8$ pixels/embryo, p<0.05). There were no differences among above concentration of antioxidants in direct comparison ($33.6{\pm}0.9{\sim}35.2{\pm}1.1$ pixels/embryo). Thus, an antioxidant of $50{\mu}M$ Vit. C was selected for SCNT. $H_2O_2$ levels of bovine SCNT embryos were significantly lower in embryos treated with Vit. C during only SCNT procedure ($26.4{\pm}1.1$ pixels/embryo, p<0.05) than the treatment group during IVM ($29.9{\pm}1.1$ pixels/embryo) and non-treated control ($34.3{\pm}1.0$ pixels/embryo). Moreover, $H_2O_2$ level of SCNT embryos treated with Vit. C during SCNT procedure was similar to that of IVF embryos. These results suggest that the antioxidant treatment during SCNT procedures can reduce the ROS generation level of SCNT bovine embryos.

• 한국수정란이식학회지
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• 제31권1호
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• pp.19-25
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• 2016

This study was conducted to evaluate the effects of type of culture media (BM, G2, OS, TCM, and MEM) on B6D2F1 mice oogenesis. In the present study, B6D2F1/CrljOri $F_1$ mice were utilized in order to maximize oogenesis. Also we used TCM-199, Dulbecco's medified Eagle's medium (DMEM), embryo culture medium (Fertilization medium, Cleavage medium, Blastocyst medium), G series medium and One step medium. In vitro maturation was highest in BM followed by the order of OS, MEM, TCM and G2 ($90{\pm}2.8%>88{\pm}3.2%>85{\pm}4.9%>78{\pm}10.2%>64{\pm}7.7%$, respectively). To note, the G2 group was statistically different compared to other groups (p<0.05). On the other hand the fertilization rate was highest in the G2 group followed by BM, OS, TCM, and MEM ($87{\pm}7.2%>85{\pm}6.9%>74{\pm}14.0%>71{\pm}13.8%>2{\pm}1.4%$, respectively). The MEM group was significantly lower compared to other groups (p<0.05). The developmental rate was highest in the OS group followed by the G2 group and the BM group albeit no statistical significance was noted ($73{\pm}11.6%>71{\pm}9.2%>66{\pm}10.4%$). Of note, all cells of the TCM and MEM groups were died during embryonic development. The zona hatched rate ($51{\pm}9.8%$ vs. $50{\pm}9.1%$ vs. $47{\pm}7.2%$ for BM, G2, and OS respectively) and attached rate ($45{\pm}12.3%$ vs. $38{\pm}16.1%$ vs. $37{\pm}11.5%$ for BM, G2, and OS respectively) were not different amongst groups. No difference was found in total cell numbers ($74{\pm}13.9$ vs. $64{\pm}9.2$ vs. $76{\pm}6.7$ for BM, G2, and OS respectively), ICM cell numbers ($20{\pm}1.9$ vs. $14{\pm}1.8$ vs. $15{\pm}2.1$), TE cell numbers ($55{\pm}12.5$ vs. $49{\pm}10.7$ vs. $61{\pm}5.9$), % ICM ($30{\pm}2.8%$ vs. $24{\pm}7.0%$ vs. $22.8{\pm}2.2%$) and ICM:TE ratio ($1:2{\pm}0.5$ vs. $1:3.1{\pm}0.8$ vs. $1:3.1{\pm}0.5$) amongst groups. In summary, these results can provide fundamental data to maximize culture condition for in vitro fertilization on B6D2F1 mice.

• Journal of Plant Biotechnology
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• 제7권4호
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• pp.259-265
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• 2005

Indian citrus ringspot virus (ICRSV)-free plants of Kinnow mandarin (Citrus nobilis Lour x C. deliciosa Tenora) were raised from virus-infected plants using unfertilised ovules as explants. Plants were tested by indirect ELISA and RT-PCR before using their explant. An amplified product of 539 bp was obtained by RT- PCR in ICRSV infected plants. Unfertilized ovules were excised from unopened flower buds of plants tested postive for virus and were cultured on Murashige and Skoog's (MS) basal medium supplemented with various concentrations of kinetin (KN) or malt extract (ME). Maximum induction (31.94%) of embryogenic callus was observed on MS medium supplemented with KN ($9.29\;{\mu}M$). Transfer of embryogenic calli to similar media composition resulted in somatic embryogenesis in all cultures, with an average number of 60.36 globular, 17.39 heart and 7.71 cotyledonary-shaped somatic embryos per culture. All cotyledonary shaped embryos developed into complete plantlets within 60 days on transfer to similar medium. Embryogenic callus induction, somatic embryo formation, maturation, germination and plantlet formation were achieved on MS medium supplemented with KN ($9.29\;{\mu}M$) alone. The plantlets derived from somatic embryos were transferred to sterilized soil, sand and vermiculite (3:1:1) mixture. After acclimatization, the plantlets were transferred to screen house and were indexed for ICRSV employing indirect ELISA and RT-PCR and found free of virus. A distinct feature of this study is the induction of somatic embryogenesis from unfertilised ovules to produce virus-free plants.

• Clinical and Experimental Reproductive Medicine
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• 제38권2호
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• pp.82-86
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• 2011

Objective: To evaluate sperm nuclear DNA fragmentation and chromatin structure after 18 hours' incubation at room temperature. Methods: Twenty-eight male partners who participating IVF treatment were prospectively included in this study. Ejaculated sperm count and motility were assessed. The sperm was then immediately processed by the conventional swim-up method. After utilization of some of the sample for routine clinical use, the remainder of each of the samples was divided into two aliquots. One aliquot was immediately assessed for sperm nuclear DNA fragmentation (TUNEL assay) and chromatin structure (toluidine blue [TB] staining). The other aliquot was incubated at room temperature for 18 hours and then assessed by two methods. Only dark-TB sperms were considered as having abnormal chromatin structure. Data before and after extended incubation were compared using a paired Student's $t$-test. Results: Before and after extended culture, nuclear DNA fragmentation assessed by TUNEL was $4.9{\pm}4.7%$ and $7.0{\pm}6.4%$, respectively ($p$=0.008). The proportion of abnormal chromatin structure (dark-TB sperm) was $8.2{\pm}5.6%$ and $10.3{\pm}6.5%$ ($p$ <0.001), before and after incubation, respectively. Conclusion: After 18 hours' incubation at room temperature, sperm nuclear DNA and chromatin structure were significantly affected. The IVF practitioner should bear this information in mind when performing delayed insemination, especially for $in$ $vitro$ maturation cycles.

• 한국동물학회지
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• 제34권2호
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• pp.181-187
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• 1991

양서류 여포를 배양할 때 외부에서 cAMP를 첨가하면 호르몬에 의한 난자의 성숙이 억제된다는 많은 보고가 있었다. 그러나 실제 외부의 cAMP가 여포내로 들어간다는 보고는 아직 없다. 본 연구에서는 배양액내의 cAMP가 여포내로 침투해 들어가는 현상과 들어간 cAMP의 분해과정을 radioimmunoassay로 조사하였다. 개구리 여포를 배양하면서 배양액에 난자의 성숙을 억제하는 농도의cAMP(2.5 mM)를 첨가한 후 일정시간 간격으로 여포내 축척된 cAMP의 농도를 조사한 결과 2시간에서 이미 기본수준(여포당 약 3 p mole)의 수십배로 증가하였다.(여포당 90 p mole). cAMP를 포함한 배양액에서 6시간 배양 후 보통 배양액으로 옮겨 배양하면서 일정 시간마다 여포내 cAMP의 농도를 측정한 결과 6시간 내에 cAMP농도가 여포당 160 p mole에서 약 10 p mole로 급격히 낮아졌다. 그러나 18시간 후에도 기본 수준으로까지 내려가지는 않았다. 이러한 cAMP의 감소과정이 progesterone이나 isobuty methylxanthine (IBMX)에 크게 영향을 받지 않았다. 배양중인 여포를 forskolin(9 u m)으로 자극했을 때에는 기본 수준의 약 3배정도로 cAMP의 농도가 증가하였다. 본 결과는 배양액내의 cAMP가 여포내로 투과해 들어가고 이들은 곧 여포에 의해 분해된다는 것을 시사하고 있다.

• 한국가축번식학회지
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• 제26권3호
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• pp.223-228
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• 2002

본 연구는 돼지의 미성숙 난포란을 체외에서 성숙, 수정시킨 후, 생산된 체외수정란을 NCSU 23 체외배양액에 항산화제NAC, ebselen 및 glutathione)와 BSO의 첨가가 돼지 체외수정란의 체외발육에 미치는 영향을 검토하였다. Glutathione 합성억제제인 BSO가 돼지수정란의 체외발육에 미치는 영향을 검토한 결과 0, 1.0 및 5.0 mM의 BSO을 첨가한 구에서 상실배기 이상 발육된 체외발육성적은 각각 35.9, 15.7 및 8.4%로서 BSO 첨가구가 대조구에 비해 통계적으로 유의하게 낮은 체외배율성적을 나타냈으며(P<0.05), NAC 1.0 mM ebselen 10 $\mu\textrm{m}$ 및 glutathione 100 $\mu\textrm{m}$ BSO 1.0 mM을 첨가하여 배양한 결과 상실배 이상 발육된 체외발육율은 40.5, 44.2, 36.0 및 10.9%로서 항산화제 첨가구가 BSO 첨가구보다 통계적으로 유의하게 높은 체외발육성적을 나타냈다P<0.05). 체외수정후 생산된 배반포기 수정란의 세포수는 BSO 농도(0, 1 및 5 mM)에 따른 차이는 인정되지 않았으나, 항산화제를 첨가한 경우에는 ebslene, NAC 및 glutathione 첨가구가 BSO 첨가구보다 유의하게 높은 세포수을 나타냈다(P<0.05).

• 한국가축번식학회지
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• 제21권3호
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• pp.303-310
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• 1997

소 난포란의 체외성숙시 성숙배지에 FSH 및 LH의 첨가가 체외성숙난자의 calcium 반응과 체외수정란의 발달에 미치는 영향을 조사하였다. 난포란의 체외성숙은 TCM199을 기초로 한 4가지의 배양조건 하에서 : 1) 0.5$\mu\textrm{g}$/ml FSH＋5$\mu\textrm{g}$/ml LH, 2) 0.5$\mu\textrm{g}$/ml FSH, 3) 5$\mu\textrm{g}$/ml LH 및 4) 무 호르몬 첨가구로서 5% CO2에 24시간 동안 체외성숙을 유도하였다. 체외성숙 24기간째에 난포란의 과립막세포는 1ml PB1＋에서 4분 동안 vortexing을 하여 완전히 제거하였다. 세포 내 calcium 반응을 측정하기 위하여 2mM Fura-2 AM ester 및 0.02% Pluronic F-127가 첨가된 PB1-용액에 39$^{\circ}C$ in cubator에서 40분 동안 배양하였다. 30${\mu}\ell$ M2 medium drop을 30mm plastic dish에 만들어 20$\times$ 형광대물렌즈가 장착된 Nikon Diaphot 현미경의 장착된 Nikon Diaphot 현미경의 warm stage에 설치하였다. 세포 내 calcium 방출을 자극하기 위하여 난자에 25mM inositol 1, 4, 5-trasphophate(IP3)로 1.21kV/cm의 전기자극 또는 20mM ryanodine으로 미세주입을 실시하였다. 이러한 처리를 하지 않은 난자는 체외수정 후 CR1aa와 BRL monolayers의 공배양조건 하에서 체외발달을 유도하였다. 분할율(Day 2)과 배반포기발달율(Day 9)을 조사하였다. FSH와 LH의 처리구에서 IP3 또는 ryanodine으로 자극된 난자(1.79$\pm$0.05, 1.66$\pm$0.06)는 FSH, LH 및 무 호르몬처리구에 비하여 유의적으로 높은 calcium 반응을 보였다(1.00$\pm$0.03, 1.28$\pm$0.04, and 0.53$\pm$0.02 in IP3 elctroporation; 0.68$\pm$0.05, 1.03$\pm$0.05, and 0.47$\pm$0.04 in ryanodine microinjection). FSH와 LH, FSH, LH처리구에서 분할율(87.9, 71.5 및 75.6%)은 무 호르몬처리구(60.7%)(P<0.05)에 비하여 유의적으로 높았으며, FSH와 LH처리구(29.3%)에서의 배반포기 발달율은 FSH, LH 처리구뿐만 아니라 무 호르몬처리구보다 유의적으로 높았다(16.5, 19.0 and 9.8%)(P<0.05). Bovine FSH 및 Ovine FSH의 처리구에서의 calcium 반응은 유의적인 차이가 없었다(1.72$\pm$0.05, 1.61$\pm$0.06). 또한 분할율(82.2 and 84.0%) 및 배반포기(27.8 and 27.1%) 발달율도 bovine 및 ovine FSH처리구간에는 유의적인 차이가 없었다. 이상의 결과에서 전기자극에 의한 세포 내 calcium 반응은 체외성숙배지에 첨가하는 호르몬의 처리에 따라서 유의적인 변화를 보였다. 비록 분할율은 처리구간에 유의적인 차이가 없었지만 배반포기 발달율은 FSH와 LH 공동처리구에서 FSH, LH 단독처리구 및 무 호르몬처리구에 비하여 유의적으로 높은 발달율을 보였다. 체외성숙기간에 FSH와 LH의 공동첨가는 체외성숙 및 체외발달의 생리적인 교정을 위하여 요구되는 것으로 사려된다.