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Annealing control primer system을 이용한 어류 재조합 myostatin prodomain 단백질에 의해 성장이 증가된 무지개송어의 특이적 발현 유전자 탐색
이상범 ( Sang Beum Lee ),진형주 ( Hyung Joo Jin ) 한국어류학회 2012 韓國魚類學會誌 Vol.24 No.2
이전 연구에서 넙치유래 재조합 마이오스타틴 프로도메인을 무지개송어에 한달간 침지법을 통하여 처리한 결과 대조군에 비하여 무게가 최대 약 42% 증가되었다. 따라서 본 연구는 재조합 마이오스타틴 프로도 메인을 침지법에 의해 처리된 무지개송어와 대조군의 근육으로부터 발현되는 cDNA를 제작하여 마이오스타틴 프로도메인에 의해서 유도된 특정유전자를 선발하기 위하여 ACP (annealing control primer)를 이용한 DDRT법 을 통하여 분석하였다. 총 20가지의 ACP를 이용한 결과 2개의 특정 유전자를 분석하였으며, NCBI BLAST분석 결과 Cytochrome P450 mono oxygenase와 Profilin으로 판명되었다. 이 중 Cytochrome P450 mono oxygenase는 대조군보다 발현량이 증가하였으며, Profilin는 대조군에 비해서 발현량이 감소하였다. 이러한 결과를 재확인하기 위하여 두 유전자의 primer를 각각 제작하여 semi-quantitative RT-PCR를 시행한 결과 DDRT법에 의한 분석과 동일하였다. 본 결과는 어류의 성장에서 마이오스타틴 프로도메인의 기능 및 메카니즘에 대한 연구에 유용한 자 료가 될 것으로 사료된다. The present study was conducted to investigate different gene expression profile between treated poMSTNpro and non-treated in rainbow trout and to identify those genes that are specifically or predominantly expressed in treated poMSTNpro by employing annealing control primer (ACP)-based GeneFishing polymerase chain reaction (PCR). We isolated total RNAs in muscle tissues from the treated poMSTNpro fish by immersion bath technique with fish myostatin prodomain (Paralichthys olivaceus, poMSTNpro) for one month and the other was non-treated poMSTNpro, and synthesized cDNA using annealing control primers (ACP, Seegene, Korea). Using 20 different ACPs for PCR, were cloned sequenced, and analyzed identities of 2 differentially expressed genes (DEGs). According to BLAST analysis, sequences of 2 clones significantly matched database entries and confirmed by semi-quantitative RTPCR. The functional roles of one up-regulated gene, cytochrome P450 mono-oxygenases 2K1v2 (CYP2K1v2), and one down-regulated gene was Profilin-1 were identified. We identified distinctive gene expression profiles in improved rainbow trout growth by treatment with a fish myostatin prodomain using ACP-based GeneFishing.
이상범(Lee, Sang-Beum),김선정(Kim, Sun-Jung) 한국체육과학회 2021 한국체육과학회지 Vol.30 No.1
The purpose of this study was to develop instructional learning framework based on the sometact method to enhance the educational effects with minimal contact. This study attempted to prove the impact of self-initiative learning ability, learning attitude, and practical capability of team based teaching through sometact for the students who are majoring in sport education. Convenience sampling method was utilized for the sample who are attending a college located in Kyounggi-Do. The interview method as a qualitative research were used to identify the effects of TBL learning by utilizing lubric likert scale of badminton. The TBL learning contents included 7 categories according to 5 evaluative criteria. To prove the effects of the method, the badminton instruction classes were conducted for 8 weeks to the sample. Findings indicated that the TBL learning was effective and students were satisfied with the sometact method. This study provide practical implications in using team based self-initiative learning with minimal contacts in the Covid-19 situation.
어류 유래 마이오스타틴 프로도메인 단백질에 의한 시마연어(Oncorhychus masou) 성장효과
Jeong Hwan Kim(김정환),Sang Beum Lee(이상범),Mi Jin Cho(조미진),Ji Young Ahn(안지영),Suk Keun Lee(이석근),Sung-Youl Hong(홍성열),Ki Baik Seong(성기백),Hyung-Joo Jin(진형주) 한국생명과학회 2011 생명과학회지 Vol.21 No.8
성장과 분화를 조절하는 인자인 myostatin은 포유류에서 주로 골격근에 분포하며 근육성장을 억제하는 것으로 알려져 있다. Myostatin은 포유류에서뿐만 아니라 어류에 있어서도 그 기능이 유사하며 본 연구에서는 넙치와 조피볼락 유래 재조합 myostatin 단백질을 생산하여 시마연어에 침지방법을 통해 처리하였다. 처리 결과 시마연어의 무게와 생화학 분석에서는 유의성이 나타날 정도의 증가는 없었지만 근(muscle) 조직학적 분석에서 넙치와 조피볼락 유래 재조합 myostatin prodomain에 의해 12주째에는 세포의 수가 증가하는 hyperplasia가 일어났으며 22주째에는 조피볼락 유래의 재조합 myostatin prodomain을 처리한 군에서만 hypertrophy가 일어났다. 결론적으로 어류 유래 재조합 myostatin prodomain이 시마연어 근육성장 시 hyperplasia와 hypertrophy가 순차적으로 유도되는 것으로 확인되었다. Myostatin (MSTN) belongs to the transforming growth factor-β superfamily or growth and differentiation factor 8 (GDF-8), and functions as a negative regulator of skeletal muscle development and growth. Previous studies in mammals have suggested that myostatin knock-out increased muscle mass and decreased fat content compared to those of the wide type. Recently, several studies on myostatin have beenconducted on the block myostatin signal pathway with myostatin antagonists and the MSTN regulation with RNAi to control myostatin function. This study was performed to analyze growth and muscle alteration of Oncorhychus masou by treatment with recombinant myostatin prodomains derived from fish. We designed myostatin prodomains derived from P. olivaceus (pMALc2x-poMSTNpro) and S. schlegeli (pMALc2x-sMSTNpro) in a pMALc2x expression vector, and then purified the recombinant proteins using affinity chromatography. The purified recombinant proteins were treated in O. masou through an immersion method. Recombinant protein treated groups did not show a significant difference in weight, protein, or lipid composition compared to the control. However, there was a difference in the average number and area for histological analyses in the muscle fiber. At twelve and twenty-two weeks from the initial treatment, there were differences in average fiber number and area between the 0.05 ㎎/ℓ treated-group and the control, but the numbers were similar to those of the control during the same time period. At twelve weeks, however, 0.2 ㎎/ℓ treated-group had an increase in average fiber number and decrease in average fiber area compared to the control. At twenty-two weeks, the pMALc2x-sMSTNpro 0.2 ㎎/ℓ treated-group was induced and showed a decrease in average fiber number and increase in average fiber area. The results between twelve and twenty-two weeks showed that the fiber numbers had decreased, whereas average fiber area had increased due to sMSTNpro. It is understood that the sMSTNpro induced only hyperplasia at twelve weeks, after which it induced hypertrophy. Recombinant myostatin prodomains derived from fish may induce hyperplasia and hypertrophy in O. masou depending upon the time that has elapsed.