Isolation and aharacterization of genes induced by a short time of treatment with Nod factors in soybean suspension-cultured cells using a PCR-based cDNA subtraction method

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  • ダイズ培養細胞にNod factorを短時間処理することで転写量が変動した遺伝子の単離
  • ダイズ バイヨウ サイボウ ニ Nod factor オ タンジカン ショリスル コト デ テンシャリョウ ガ ヘンドウシタ イデンシ ノ タンリ

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Abstract

Lipochito-oligosaccharides (Nod factors) produced by Rhizobium or Bradyrhizobium are key signaling molecules that elicit nodulation in their corresponding host legumes. To clarify gene expression induced by treatment with Nod factors in soybean suspension-cultured cells, we subtracted cDNAs induced by the Nod factors from those induced by elicitors of Mycosphaerella pinodes using a PCR-based cDNA subtraction method. We isolated four cDNA clones: nsc1 and nsc2 had high DNA homology to soybean expressed sequence tags (ESTs) obtained from super-nod plants inoculated with Bradyrhizobium japonicum USDA110 prior to harvest, nsc3 had high DNA homology to a soybean 4-Coumarate: CoA ligase (4CL) gene, and nsc4 had high similarity to a soybean EST obtained from 2- to 3-week-old soybean seedlings in which sudden death syndrome had been induced by Fusarium solani. We reconfirmed expression of genes corresponding to the nsc clones induced by Nod factors in soybean suspension-cultured cells, but no gene expression was induced by M. pinodes elicitors. To elucidate changes in gene expression profiles over time, we monitored levels of transcripts corresponding to the nsc clones in the suspension-cultured cells with the Nod factors for 4h using the RT-PCR/Southern method. Levels of all transcripts increased rapidly within 10min after treatment with the Nod factors, then gradually decreased, except for nsc4, whose transcripts remained stable. For nsc3, a homologue of soybean 4CL was isolated from soybean roots, and changes over time in its transcription level in soybean germinated roots after treatment with Nod factors were monitored using RT-PCR. We also confirmed that these roots showed a more rapid increase in 4CL transcripts than in roots without the Nod treatment.

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