• Journal of Photoscience
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• v.9 no.2
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• pp.278-280
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• 2002

We have cloned a hydromedusan opsin cDNA and showed that the deduced amino acid sequence of the cytoplasmic loop between helices 5 and 6 (loop 5-6) was clearly different from that reported so far. The amino acid sequence of the loop 5-6 is important on determination of the specificity for the coupled G- protein. To clarify which class of G-protein mediates the phototransduction system in the ocellus of the hydromedusan, we investigated G-proteins expressed in the ocellus. By PCR against the cDNA of the ocellus with primers designed according to the conserved amino acid sequence in G-protein a subunit, we obtained three kinds of cDNA fragments. Based on the sequence similarities, ttwo of them (JGI and JG3) were classified as $G_{i}$ and $G_{q}$, respectively. The other one (JG2) was a new subtype within $G_{*}$ class. Electron microscopic immunocytochemistry with the antiserum against the C-terminal sequence of $G_{q}$ or $G_{t}$ revealed the presence. of the both classes in the ocellus. The similarity of the C-terminal sequence of the JG2 with that of bovine $G_{t}$ suggests that the anti- $G_{t}$ antiserum would bind to JG2. These results suggest the possibility that the hydromedusan rhodopsin decides the specificity for the coupled G-protein by the other domain than the loop 5-6.oop 5-6.5-6.

• Korean Journal of Animal Reproduction
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• v.22 no.1
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• pp.1-9
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• 1998

This study was carried out to confirm whether the developmental capacity of bovine mature oocytes frozen ultra-rapidly using electron microscopic(EM) grids and EFS30 can be obtained, and whether the cryoprotectants and the freezing method used in this study effect detrimentally to the bovine oocytes by indirect immunocytochemistry. As freezing solution, we used EFS30 which consisted of 30% ethylene glycol, 0.5 M sucrose, 18% ficoll and 10% FBS added in D-PBS. The results obtained in this experiment were summarized as follows: When the effects of cryoprotectant and freezing procedure on the microtuble, micrfilament and chromatin morphology of oocytes were evaluated using indirect immunocytochemistry, the results of freezing as well as exposure group were not different with that of the control oocytes. When the fertilization abnormality after ultrarapid freezing of bovine mature oocytes was examined by Hoechst staining, the rates of total penetration(96.7, 9.0%), normal two pronuclei formation(74.6, 68.9%) and mean number of sperm / oocyte(1.50, 1.44) were not different between control and freezing group. In addition, when the developmental capacity of frozen-thawed of oocytes(85.5%) was survived, 74.5% of them were cleaved and 31.4% of cleaved embryos were developed to blastocyst. These data were similar to those of the control(76.0%, 34.6%) and exposure(74.5%, 33.0%) except survival rates. Also, when the total cell number of blastocysts produced from the each treatment at day after IVF was examined by hoechst staining, there were not different among groups. There results demonstrate that developmental capacity of frozen-thawed bovine mature oocytes can be successfully obtained by ultra-rapid freezing method using EM grid and EFS30 solution.