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Gene expression profile of the early embryonic gene of the silkworm, Bombyx mori

누에 수정란 초기발현유전자 데이터베이스 구축

  • Choi, Kwang-Ho (Sericultural & Apicultural Materials Division, National Academy of Agricultural Science, RDA) ;
  • Goo, Tae-Won (Sericultural & Apicultural Materials Division, National Academy of Agricultural Science, RDA) ;
  • Kim, Seong-Ryul (Sericultural & Apicultural Materials Division, National Academy of Agricultural Science, RDA) ;
  • Kim, Sung-Wan (Sericultural & Apicultural Materials Division, National Academy of Agricultural Science, RDA) ;
  • Chun, Jae-Buhm (Sericultural & Apicultural Materials Division, National Academy of Agricultural Science, RDA) ;
  • Park, Seoung-Won (Dep. of Biotechnology, Catholic Univ. of DAEGU) ;
  • Kang, Seok-Woo (Sericultural & Apicultural Materials Division, National Academy of Agricultural Science, RDA)
  • 최광호 (농촌진흥청 국립농업과학원 잠사양봉소재과) ;
  • 구태원 (농촌진흥청 국립농업과학원 잠사양봉소재과) ;
  • 김성렬 (농촌진흥청 국립농업과학원 잠사양봉소재과) ;
  • 김성완 (농촌진흥청 국립농업과학원 잠사양봉소재과) ;
  • 전재범 (농촌진흥청 국립농업과학원 잠사양봉소재과) ;
  • 박승원 (대구가톨릭대학교 의료생명산업대학 생명공학과) ;
  • 강석우 (농촌진흥청 국립농업과학원 잠사양봉소재과)
  • Received : 2013.09.28
  • Accepted : 2013.10.15
  • Published : 2013.11.30

Abstract

This study was aimed for development of a useful genes that has a transcript expressional specificity in the early embryonic stage of the silkworm, Bombyx mori. We constructed and analyzed a full-length cDNA library from silkworm's eggs which after a lapse of 2 ~ 6 hours post oviposit. A total 960 clones were randomly selected, and the 5' ends of the inserts were sequenced to generate 652 expressed sequence tags(EST). 334 unique ESTs were generated after the assembly of 652 ESTs. The annotation of 334 unique ESTs by BLAST search revealed that 156(47%) of the sequences represented known genes, whereas 178(53%) of the sequences has no matches in the database. Of the 156 known genes, the most abundant genes were heat shock protein hsp20.8 gene(12 times) and ubiqutin-like protein gene(11 times). The functional groups of these ESTs with matches in the database were constructed according to their putative molecular functions. Among thirteen functional categories, the largest groups were protein synthesis(9.6%) and cellular organization( 8.1%). Further defined studies on molecular functions and biological roles of their promoters will give us wellfined information and its application.

본 연구는 누에 수정란 초기에 발현하는 유전자를 대량 선발하고, 유용 유전자의 프로모터를 개발하기 위한 연구의 일환으로 추진하였다. 산란 후 2 ~ 16시간이 경과한 누에알로부터 cDNA 유전자은행을 제작하였다. 제작된 cDNA 유전자은행으로 전체 960개 클론을 무작위 추출하여 부분 염기서열 분석을 통해 EST를 제작하였다. 분석된 652개 ESTs 중 염기서열 상동성 분석을 통해 156개의 기존 알려진 유전자와 178개의 미지의 유전자로 구성된 334개 독립유전자를 최종 선발하여 'eegEST'로 명명하였다. eegEST 분석 결과, 기존 염기서열 정보가 알려진 156개 독립유전자 중 2회 이상 출현한 유전자 수는 143개로 전체의 34%를 차지하였으며, Hsp20.8 유전자(12회)와 ubiqutin-like 유전자(11회)가 가장 높은 출현 빈도를 나타내었다. 또한 eegEST 독립유전자의 추정 기능에 따른 분류에서 곤충 수정란 발생초기에 확인할 수 있는 기관 형성과 관련한 유전자가 전체 24%를 차지하고 있었다. 본 연구에서 작성된 누에 수정란 초기 발현유전자 데이터베이스(eegEST)는 곤충 발생학 연구를 위한 정보제공 뿐 아니라 형질전환누에 제작을 위한 프로모터 개발 연구에 활용될 수 있을 것으로 기대한다.

Keywords

References

  1. Ashburner M, Ball CA, Blake JA, Botstein D, Butler H, Cherry JM, Davis AP, Dolinski K, Dwight SS, Eppig JT, Harris MA, Hill DP, Lsseltarver L, Lasarskis A, Lewis S, Matese JC, Richadson JE, Ringwald M, Rubin GM, Sherlock G (2000) Gene ontology; tool for the unification of biology. Nat Genet 25, 2529.
  2. Boo KS, Kim YG, Park GC, Choi MY (2005) Embryogenesis; in Insect hormone and physiology, pp.645651, Seoul national university Press, Seoul.(in Korea).
  3. Choi KH, Goo TW, Yun EY, Hwang JS, Kang SW (2007) Gene expression profile of the posterior silk glands of the silkworm, Bombyx mori L., Korean J Genetics 29(2), 2-9.
  4. Goldsmith MR, Shimada T, Abe H (2005) The genetics and genomics of the silkworm, Bombyx mori. Annu Rev Entomol 50, 71-100. https://doi.org/10.1146/annurev.ento.50.071803.130456
  5. Guo TQ, Wang JY, Guo XT, Wang SP, Lu CD (2005) Transient in vivo gene delivery to the silkworm Bombyx mori by EGT-null recombinant AcNPV using EGFP as a reporter. Arch Virol 150, 93-105. https://doi.org/10.1007/s00705-004-0383-y
  6. Handler AM (2001) A current perspective on insect gene transformation. Insect Biochem Mol Biol 31, 111-128. https://doi.org/10.1016/S0965-1748(00)00159-4
  7. Hwang JS, Go HJ, Goo TW, Seong SI, Yun EY, Ahn MY, Kim SR, Park KH, Kim IS, Jeon JP, Kang SW (2007) Molecular characterization of small heat shock protein(hsp20.8A) from the silkworm, Bombyx mori. Int J Indust Entomol 15(1), 75-78.
  8. Imamura M, Nakai J, Inoue S, Guo XQ, Kanda T, Tamura T (2003) Target gene expression using the GAL4/UAS system in the silkworm Bombyx mori. Genetics 165, 1329-1340.
  9. Kim SW, Yun EY, Choi KH, Kim SR, Park SW, Kang SW, Kwon OY, Goo TW (2012) Construction of fluorescent red silk using fibroin H-chain expression system. J Seric Entomol Sci 50(2), 87-92. https://doi.org/10.7852/jses.2012.50.2.87
  10. Kim SW, Yun EY, Choi KH, Kim SR, Kang SW, Goo TW (2013) Utilization of the Bombyx mori heat shock protein 70 promoter for screening transgenic silkworm. Entomological Research 43(5), 282-287. https://doi.org/10.1111/1748-5967.12031
  11. Lee SH, Kim BG, Kim KJ, Lee JS, Yun DW, Hahn JH, Kim GH, Lee KH, Suh DS, Kwon ST, Lee CS, Yoo YB (2002) Comparative analysis of sequences expressed during the liquid-cultured mycelia and fruit body stage of Pleurotus ostreatus. Fungal Genetics and Biology 35, 115135.
  12. Mita K, Kasahara M, Sasaki S, Nagayasu Y, Yamada T, Kanamori H, Namiki N, Kitagawa M, Yamashita H, Yasukochi Y, Okuda KK, Yamamoto K, Ajimura M, Ravikumar G, Shimomura M, Nagamura Y, Shin-I T, Abe H, Shimada T, Morishita S, Sasaki T (2004) The genome sequence of silkworm, Bombyx mori. DNA Res 29, 27-35.
  13. Mita K, Morimyo M, Okano K, Koike Y, Nohata J, Kawasaki H, Kuda KK, Yamamoto K, Suzuki MG, Shimada T, Goldsmith MR, Maeda S (2003) The construction of an EST database for Bombyx mori and its application. Proc Natl Acad Sci 100, 14121-14126. https://doi.org/10.1073/pnas.2234984100
  14. Olsen M, Hood L, Cantor C, Botstein D (1989) A common language for physical mapping of the human genome. Science 245, 1434-1435. https://doi.org/10.1126/science.2781285
  15. Ote M, Mita K, Kawasaki H, Seki M, Nohata J, Kobayashi M, Shimada T (2004) Microarray analysis of gene expression profiles in wing discs of Bombyx mori during pupal ecdysis. Insect Biochem Mol Biol 34, 775-784. https://doi.org/10.1016/j.ibmb.2004.04.002
  16. Park SW, Choi KW, Goo TW, Kim SR, Kang SW (2011) Characterization of the promoter controling the stage-specific gene expression of Bombyx mori. Journal of Life Science 21(10), 14661472. https://doi.org/10.5352/JLS.2011.21.10.1466
  17. Ponnuvel KM, Murthy GN, Awasthi AK, Rao G, Vijayaprakash NB (2010) Differential gene expression during early embryonic development in diapause and non-diapause eggs of multivoltine silkworm Bombyx mori. Indian J Exp Biol 48, 11431151.
  18. Rika H, Tomita M, Yoshzato K (2006) The generation of germline transgenic silkworms for the production of biologically active recombinant fusion proteins of fibroin and human basic fibroblast growth factor. Biomaterials 27, 5715-5724. https://doi.org/10.1016/j.biomaterials.2006.07.028
  19. Satoshi Y, Zhu Z, Iaso K, Uchino K, Tamada Y, Tamura T, Asakura T (2007) Improving cell-adhesive properties of recombinant Bombyx mori silk by incorporation of collagen or fibronectin derived peptides produced by transgenic silkworms. Biomacromolecules 8, 3487-3492. https://doi.org/10.1021/bm700646f
  20. Tamura T, Thibert T, Royer C, Kanda T, Eappen A, Kamba M, Komoto N, Thomas JL, Mauchamp B, Chavancy G, Shirk P, Fraser M, Prudhomme JC, Couble P (2000). A piggyBac element-deribed vector efficiently promoters germ-line transformation in the silkworm, Bombyx mori L. Nat Biotechnol 18, 81-84. https://doi.org/10.1038/71978
  21. Tomita M (2011) Transgenic silkworms that weave recombinant proteins into silk cocoons. Biotechnol Lett 33, 645-654. https://doi.org/10.1007/s10529-010-0498-z
  22. Velu D, Ponnuvel KM, Syed MHQ (2008) Expression of the heat shock protein genes in response to thermal stress in the silkworm Bombyx mori. Int J Indust Entomol 16(1), 2127.
  23. White LD, Coares CJ, Atkinson PW, O'Brochta DA (1996) An eye color gene for the detection of transgenic non-drosophilid insects. Insect Biochem Mol Biol 26, 641-644. https://doi.org/10.1016/S0965-1748(96)00039-2