• 한국수정란이식학회지
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• 제13권2호
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• pp.139-145
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• 1998

In the present study, effects of interleukin-2 (IL-2), a differentiator and proliferator of T-cells, on nuclear maturation and sperm penetration of bovine oocytes was examined in a serum-free or serum-containing medium. Basic medium was used TCM-199 supplemented with 2.2g / ι sodium bicarbonate, 100 i.u. /rnl penicillin. 100$\mu$g /ml streptomycin, 0.25$\mu$g/ml Fungizone, this medium treated with FCS and IL-2. In experiment 1, we examined the effect of the addition of 0, 1, 5, 10 or 15nM /ml IL-2 to tissue culture medium (TCM-199) on nuclear maturation of oocytes Development of oocytes to the Metaphase II (M II) stage (%) was significantly (P<0.05) higher at 1, 5,10 and 15 nM /ml IL-2(54.2, 73.5, 80.0 and 69.6%, respectively) than at 0 nM /ml IL-2(35.7%). In experiment 2, we examined the effect of the addition of l0nM /ml IL-2 or 5% FCS in oocyte maturation. Nuclear maturation rates were significantly(P<0.05) higher l0nM /ml IL-2(80%) than non-treatment(35.7%) and 5% FCS(63.6%) treatment. On the other hand, there were no significant difference in the proportion of oocytes developed to the 2-cell stage after addition of IL-2 and/or FCS. These results suggest that IL-2 supports nuclear maturation of bovine immature oocytes in vitro. Serum-free maturation system using IL-2 might be useful for evaluation of various factors on oocyte maturation.

• 한국수정란이식학회지
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• 제3권1호
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• pp.13-23
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• 1988

These studies were undertaken to examine the interaction of tamoxifen with sex steroid hormones in rat uterine activity. The uterine wet weights of the immature Tat uterus were examined after the administration of estradiol-l7$\beta$(1$\mu$g), tamoxifen(50$\mu$g), progesterone(lmg). The uterotropic activity in immature ovariectomized rats was observed under various treatment conditions following pretreatment with above drugs. The results obtained were as follows:1) Tamoxifen produced significant increase (p <0.01) in uterine wet weight compared with control group, although the increase was not as great as that seen with estradiol-17$\beta$. Administration of estradiol-17$\beta$ together with tamoxifen inhibited significantly the increase of uterine wet weight by estradiol-17$\beta$ (p < 0.01). Coadministration of progresterone with tamoxifen partly blocked the increase of tamoxifen-induced uterine wet weights by progesterone. 2) Estradiol-17$\beta$after the estradiol-17$\beta$ pretreatment discontinued the declining uterine wet weights due to the absence of estrogen support, but uteri continued to increase in weight if daily estradiol-17 $\beta$ was maintained. Administration of tamoxifen on the fourth day of estradiol-17$\beta$ treatment reduced uterine wet weights within 24 hours, and the weights continued to decline with additional tamoxifen. 3) The modest growth of the uterus induced by three daily injections of 5Opg tamoxifen remained stable for five days, with or without additional tamoxifen treatment. Coadministration of tamoxifen with estradiol17$\beta$ increased slightly the increase of uterine wet weight by tamoxifen. Coadministration of tamoxifen with progesterone inhibited the increase of uterine wet weight by tamoxifen. 4) The modest growth of the uterus induced by three daily injections of lmg progesterone reduced uterine wet weight to the control level for five days. Commencement of tamoxifen or estadiol-17 $\beta$ injections on the fourth day of progesterone treatment rapidly elevated uterine wet weight.

• 한국수정란이식학회지
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• 제24권2호
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• pp.121-125
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• 2009

In the present study, effects of concentration of cryoprotectant solutions on the nuclear maturation of vitrifiedthawed bovine oocytes were examined. Also, the developmental capacity of vitrified-thawed immature oocytes following ICSI was investigated. Oocytes were cultured in TCM-199 medium supplemented with 5% FBS at $38^{\circ}$C in 5% $CO_2$ and air. The in vitro maturation rate of vitrified oocytes was 24.5 ${\pm}$ 4.2%. The in vitro maturation rate of vitrified oocytes was lower than that of the control (72.0 ${\pm}$ 3.5%, p<0.05). The in vitro maturation rate of vitrified${\sim}$thawed oocytes incubated in TCM-199 medium supplemented with 1.0${\sim}$5.0 ug CB were 26.7 ${\pm}$ 3.2%, 35.7 ${\pm}$ 3.2%, 54.0 ${\pm}$ 3.0%, 42.5 ${\pm}$ 3.6%, respectively. The in vitro maturation rate (57.0 ${\pm}$ 3.0%) of the vitrified-thawed oocytes treated with 3.0 ${\mu}$g CB for 20 min was the highest of all vitrification groups, although the maturation rate were significantly (p<0.05) lower than those of fresh oocytes. The in vitro maturation rates of the vitrified-thawed (with EDS and EDT) oocytes were 53.8 ${\pm}$ 3.4%, 51.1 ${\pm}$ 3.5%, respectively. This results were lower than the control group (72.0 ${\pm}$ 3.0%). The in vitro developmental rates of the vitrified-thawed oocytes following ICSI were 28.6 ${\pm}$ 4.5%, 25.6 ${\pm}$ 4.3%, respectively. This results were lower than the control group (40.0 ${\pm}$ 4.0%).

• Journal of Plant Biotechnology
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• 제32권2호
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• pp.91-95
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• 2005

옥수수 미숙배배양과 Agrobacterium tumefaciens공동배양법에 의해 형질전환체를 생산하였다. Hi II계통의 미숙배를 Ubiquitin 1 promoter-GUS유전자와 선발마커로서 nptII 유전자로 제작된 pPTN290벡터를 C58C1에 도입한 후 형질전환 균주로 사용하였다. 7개의 paromomycin저항성 배 발생캘러스를 얻었으며, GUS양성반응을 나타내는 7개의 독립적인 식물체를 얻었다. Southern분석법에 의하여 $T_1$세대 식물체로부터 nptII유전자가 안정적으로 도입되어 있음을 확인하였다.

• 식물조직배양학회지
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• 제22권4호
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• pp.189-194
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• 1995

유자의 배배양에 의한 캘러스의 유도는 미숙열매의 배로부터 가장 양호하게 유기되었다. 이들 캘러스는 3% sucrose 와 44.4 $\mu$M BA가 첨가된 1/2 MS배지에 치상하였을 때 가장 잘 유도되었으며 배양조건은 26$^{\circ}C$ 에서 2000 lx로 16 h의 광주기와 8 h의 암주기로 배양하였다. 이들 캘러스는 담황색으로 표면에 크고 작은 많은 돌기들을 가지고 있는 캘러스와 암갈색의 캘러스가 관찰되었다. 또한 캘러스 발달 단계와 기관분화의 정도에 따라서 세포배열 양상에 많은 차이가 있었는데, 배양 6주된 캘러스는 작은 세포들이 조밀하게 나타났으며 8주경부터 신장된 세포들이 분열하는 양상을 보여 주었으며 주된 캘러스 표면의 돌기들은 기관 발생의 초기에 구형의 세포돌기가 융기된 반면 주변의 세포들 은 액포화된 것으로 나타났다. 배양 16주 후에, 재분화된 shoot는 MS 배지에 이식하였을 때 발근이 되어 정상적으로 생장하였다. Shoo떼서는 유관속 형성층과 정유를 함유하고 있는 유낭의 초기 발생과정이 광학현미경으로 관찰되었다. 형성된 shoot를 화분에 이식하였을때 정상적으로 생장이 계속되는 것을 관찰하였다.

• 한국작물학회지
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• 제37권3호
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• pp.257-263
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• 1992

GA$_3$ 농도 및 저온처리 기간이 보리의 춘화처리효과에 미치는 영향을 조사하고자, 추파성 정도가 IV인 올보리를 공시하여 출수후 13 및 20일이 경과한 미숙배를 GA$_3$농도별로 B$_{5}$배지에 치상하여 저온처리한 후 온도 20/15$^{\circ}C$, 14시간 일장으로 온냉조절실에 이식재배하여 실험한 결과는 다음과 같다. 1. 미숙배의 생육은 출수후 13일배 보다 20일배가 양호하여 발아율 및 유묘장은 증가하였고, 평균발아일수는 감소하였는데, GA$_3$1 및 10ppm이 효과가 좋았다. 2. GA$_3$농도가 증가할수록 상엽전개 및 출수일수가 단축되었으며, 춘화처리에 효과가 높은 GA$_3$농도는 1,10ppm이었고 GA$_3$와 저온처리가 병행될 때 춘화처리효과가 높았다. 3. GA$_3$ 처리에 따른 상엽전개일수 단축 효과는 3주 저온처리의 GA$_3$무처리구보다 2주 저온처리의 GA$_3$1,10ppm에서 각각 3, 16일 단축되었고, 3주저온처리의 1ppm에서 18일, 10ppm에서 20일 단축되었다. 4. 간장, 수장 및 일수립수는 공히 3주 저농처리에 GA$_3$ 10ppm 처리가 최소치를 보였는데, 실제육종상 중요한 일수립수의 확보는 충분할 것으로 보였다. 5. GA$_3$농도와 주요 형질과의 상관은 1주 저온처리시 각 형질들과 모두 상관 관계가 인정되지 않았으나, 2주 저온처리에서는 고도의 부상관이 인정되었고, 3주 저온처리에서는 출수일수를 제외한 모든 형질이 부의 상관이 있었다. 6. 저온처리 기간과 주요 형질들과의 상관은 GA$_3$무처리시 간장만 유의적인 부의 상관이 있었고, 기지 상관이 없었으며 GA$_3$1, 10ppm 처리시 모든 형질이 고도의 부상관이 있었다.다.

• 한국자원식물학회지
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• 제37권3호
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• pp.263-269
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• 2024

Embryos of early-ripening peaches could not achieve physiological maturation or undergo abortion before harvest. Embryo rescue is an effective strategy to rescue embryos from early-ripening peaches. Thus, the current study was carried out to determine the appropriate developmental stage and optimal medium composition for embryo rescue in early-ripening peach. Development of open-pollinated 'Yumi' fruit was investigated from 20 to 90 days after full bloom (DAFB) to explore period occurring endocarp hardening. After endocarp hardening, embryo development was observed by light microscopes. Shoot and root meristems were observed at 65 DAFB and embryo size rapidly increased at 75 DAFB. Embryos collected at 75, 80, 85, and 90 DAFB were cultured on four media based on Driver and Kuniyuki (DKW) medium. Germination rate of embryos cultured on four media gradually increased from 75 to 90 DAFB and reached 100% at 90 DAFB. Notably, M3 medium (0.5 DKW supplemented with 6-benzylaminopurine (BAP) 1.0 ㎎/L) displayed the highest germination rate at 75 and 80 DAFB stages. Growth and development of shoot and root were pronounced in plantlet cultured at 90 DAFB stage. While delayed shoot growth was evident in plantlets cultured at 75, 80, and 85 DAFB stages, this retardation could be overcome through the application of growth regulators, particularly in M3 and M4 (0.5 DKW supplemented with BAP 1.0 ㎎/L and indole-3-butyric acid 0.5 ㎎/L) media. Remarkably, roots of plantlet grown in M4 medium exhibited limited elongation. In conclusion, germination rate of embryo and growth of embryo cultured plantlet can be enhanced by collecting seeds from early-ripening 'Yumi' at the 90 DAFB stage and conducting embryo culture using the M3 medium.

• 한국수정란이식학회지
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• 제24권2호
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• pp.97-104
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• 2009

The objective of this study was to examine the effect of macromolecule in a maturation medium on nuclear maturation, intracellular glutathione (GSH) level of oocytes, and embryonic development after parthenogenetic activation (PA) and somatic cell nuclear transfer (SCNT) in pigs. Immature pig oocytes were cultured in maturation medium that was supplemented with each polyvinyl alcohol (PVA), pig follicular fluid (pFF) or newborn calf serum (NBCS) during the first 22 h and the second 22 h. Oocyte maturation was not influenced by the source of macromolecules during in vitro maturation (IVM). Embryo cleavage and cell number in blastocyst after PA was altered by the source of macromolecule but no difference was observed in blastocyst formation among treatments. Oocytes matured in PVA-PVA medium showed lower rates of oocyte-cell fusion (70.4% vs. 77${\sim}$82%) and embryo cleavage (75% vs. 86${\sim}$90%) after SCNT than those matured in other media but blastocyst formation was not altered (13${\sim}$27%) by different macromolecules. pFF added to IVM medium significantly increased the intracellular GSH level of oocytes compared to PVA and NBCS, particularly when pFF was supplemented during the first 22 h of IVM. Our results demonstrate that source of macromolecule in IVM medium influences developmental competence of oocytes after PA and SCNT, and that pFF supplementation during the early period (first 22 h) of IVM increases intracellular GSH level of oocytes.

• 한국수정란이식학회지
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• 제12권3호
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• pp.283-291
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• 1997

In vitro development of bovine embryos is affected by many factors such as energy substrates, amino acids, and some growth factors. It has been reported that mRNA of insulin, PDGF and their receptors are detected in cow embryos, and that some chelating agents such as EDTA and transferrin have beneficial role on mouse and bovine embryos. The author hypothesized that insulin, transferrin arid PDGF added to a culture medium increase in vitro development of bovine embryos by chelating toxic substance(s) or increasing cell growth and metabolism. Immature oocytes from slaughtered ovaries of Holstein cows and heifers were matured for 24 hours in a TCM199 containing 10% fetal calf serum, FSH, LH and estradiol with granulosa cells in vitro. Matured oocytes were coincubated with sperm for 30 hours in a modified Tyrode's medium (IVF). Embryos cleaved to 2- to 4-cell at 30 hours after IVF were selected and cultured in a 30-$\mu$l drop of a synthetic oviduct fluid medium (SOFM) containing 0.8% BSA, Minimum Essential Medium essential and non-essential amino acids, and insulin, transferrin or PDGF for 9 days. Supplementation of a SOFM with insulin, and /or transferrin did not increase develop-mental rate to expanding and hatching blastocyst of 2- to 4-cell bovine embryos compared with control. The highest developmental rate to hatching blastocyst was shown when PDGF was added at the concentration of 10 ng /ml among the supplementing doses tested in the present study (p<0.05). Addition of PDGF without insulin to a SOFM could not increase embrye development, but combined addition of PDGF with insulin significantly increased (p<0.05) embryo development to hatching blastocyst (50%) compared with control (38%). In conclusion, insulin and PDGF supplemented to a SOFM may act synergistically and have beneficial effect on in vitro development of 2- to 4-cell bovine embryos matured and fertilized in vitro.

• Journal of Plant Biotechnology
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• 제42권4호
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• pp.388-395
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• 2015

본 연구는 약용 및 식용으로 유망한 음나무의 대량생산 및 생식질 보존을 위한 기초 자료를 제공하기 위해 배발생 조직을 재료로 저온저장 기간별 SE 유도 및 식물체 재생을 시험하였다. 배발생 캘러스는 저온저장 6개월까지 정상적인 체세포배 형성이 가능하였으나 유도 빈도는 저장 기간에 따라 감소하는 경향을 보였다. 배발생능이 가장 좋은 ZE 10 세포주은 체세포배 유도 빈도에 있어 저장 기간에 따른 감소폭이 비교적 적은 것으로 나타났다. 체세포배의 발아 및 식물체 재생은 캘러스의 저장기간에 따라 다소 감소하는 경향을 보였으나 발아율은 93% 이상, 식물체 재생율은 91% 이상으로 저온저장에 따른 큰 문제는 없었다. 조직검경 결과 저장 기간에 따라 세포의 갈변화 및 활력 저하가 나타났으며, 아세토카민과 에반스블루의 염색을 통해서도 확인되었다. 이상의 결과로 볼 때 음나무 배발샐 캘러스의 저온저장은 계대배양 없이 $4^{\circ}C$의 냉장고에서 6개월 까지 가능한 것으로 나타났으며, 저장 후 체세포배 유도효율을 위해 배발생능이 양호한 세포주의 저장이 필요한 것으로 관찰되었다.