報告書番号

U95035

タイトル（和文）

分子育種法による高鉄含量レタスの作出（その２）－タバコを用いたフェリチン遺伝子導入効果の確認－

タイトル（英文）

BREEDING OF HIGH-IRON CONTENT IETTUCE (LACTUCA SATIVA L.) (2)EXPRESSION OF INTRODUCED FERRITIN GENE AND IRON ACCUMULATION IN TRANSGENIC TOBACCO PLANTS

概要 （図表や脚注は「報告書全文」に掲載しております）

高鉄含量レタスの分子育種を目的として研究を開始した。前報において，レタスとダイズのフェリチン遺伝子の発現の違いを明らかにすると共に，ダイズフェリチン遺伝子をレタスへ導入する技術を確立した。本報告では，導入した遺伝子の機能を詳細に解析するために，実験植物であるタバコを材料として，外来遺伝子の発現・翻訳の解析，および，植物体中の鉄含量の変化を測定した。その結果，遺伝子が導入されたことが確認された植物において，フェリチン遺伝子が常に発現していることが明らかとなった。また，外来遺伝子由来と考えられているタンパク質も検出された。さらに植物体中の鉄含量は，非形質転換体の１．２倍という高い値を示した。以上のことから，フェリチン遺伝子を導入することにより，高鉄含量レタスの新品種作出の可能性を示すことができた。

概要 （英文）

LETTUCE (LACTUCA SATIVA L.) IS A POPULAR VEGETABLE CROP SPECIES IN VEGETABLE FACTORIES DEVELOPED RECENTLY FOR CONTINUOUS PRODUCTION OF VEGETABLES BY CONTROLLING AIR AND USING ARTIFICIAL LIGHT. IN A PREVIOUS REPORT,WE DEMONSTRATED THE POSSIBILITY OF BREEDINGOF NEW-TYPE LETTUCE FOR VEGETABLE FACTORIES BY GENE MANIPULATION. THE SOYBEAN (GLYCINE MAX MERR) FERRITIN GENE WAS INTRODUCED INTO LETTUCES USING THE 35S PROMOTER FROM CAULIFLOWER MOSAIC VIRUS. THE IRON-STORAGE PROTEIN,FERRITIN,IS WIDELY DISTRIBUTED IN LIVING ORGANISMS. IN ANIMALS,IT IS FOUND IN A CYTOPLASMIC PROTEIN IN MANY CELL TYPE. HOWEVER,IN HIGHER PLANTS IT IS FOUND IN CHLOROPLASTS AND OTHER PLASTIDS AND NOT IN CYTOPLASM. IT IS BELIEVED THAT FERRITIN IN HIGHER PLANTS PLAYS AN IMPORTANTROLE FOR IRON ACCUMULATION AND PROTECTION OF CELLS AGAINST THE TOXIC EFFECTS OF IRON OVERLOAD. IN THIS STUDY,WE USED TRANSGENIC TOBACCO (NICOTIANA TABACUM L.) PLANTS TO INVESTIGATE IN WHICH TISSUE FERRITIN IS SYNTHESIZED IN THE PLANT AND HOW MUCH IS INCREASED IN THEIR LEAVES. TOBACCO IS MORE USEFUL THAN LETTUCE FOR SOME REASONS:THE GROWTH PERIOD OF LETTUCE IS LONGER THAN TOBACCO,AND TOBACCO IS A WELL-STUDIED PLANT INCLUDING TRANSFORMATION METHODS. TOTAL PROTEIN WAS EXTRACTED FROM LEAVES OF TRANSGENIC TOBACCO PLANTS CONTAINING SOYBEAN FERRITIN GENE AND ANALYSED BY SDS-PAGE. IMMUNOBLOTS WERE CARRIED OUT USING ANTISERUM RAISED AGAINST SOYBEAN FERRITIN PEPTIDE DERIVED FROM ESCHERICHIA COLI CONTAINING SOYBEAN FERRITIN GENE WITH THE EXPRESSION VECTOR. IMMUNODETECTION SHOWED THAT THE FERRITIN SUBUNIT EXISTED IN TRANSFORMANTS CONSTITUTIVELY,BUT NOT IN CONTROL PLANTS. THERE ARE SOME DIFFERENCES IN MOLECULAR WEIGHTS BETWEEN FERRITIN FROM TRANSFORMANTS AND FROM E. COLI,SUGGESTING THE FERRITIN SUBUNIT WAS MODIFIED IN PLASTIDS. THIS WOULD MEAN THAT THE FERRITINDERIVED FROM THE EXOGENOUS GENE PROBABLY HAS A FUNCTION FOR ACCUMULATING IRON. FURTHEMORE,THE IRON CONTENT IN THE DRY MATTER OF LEAVES OF TRANSFORMANTS WAS 20% HIGHER THAN THAT OF

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