綠竹與麻竹白化苗細胞及分子層次之檢測

Abstract

以竹類組織培養方法繁殖竹苗可供無病毒建康苗株，但在組織培養增殖過程中常出現白化苗。許多禾本科植物利用小孢子體培養所產生的白化苗成因與葉綠體基因組缺失有關，但以芽體增殖所產生的白化苗其發生之致因卻尚未有相關的發表。本研究利用芽體增殖的麻竹與綠竹白化苗為材料並利用電子顯微鏡技術、聚合酶連鎖反應、反轉錄聚合酶連鎖反應及生物晶片技術進行與正常型的比較。其中在麻竹(Dendrocalamus Iatiflorus Munro) 白化苗葉綠體基因組篩選出8個重複數下降的基因，其中白化苗中psaB (PSI P700 apoprotein A2) 的DNA重覆數只有正常型的17%。綠竹 (Bambusa oldhamii) 三種不同繼代時間所得之白化苗 (ow1、ow2及ow3)，型態上隨著繼代的時間增加，平均株高逐漸下降。在細胞結構方面，相較於正常型細胞具有結構完整的葉綠體，三種白化苗細胞只能觀察到色體 (plastid) 存在。利用丙酮萃取檢測綠竹正常型及白化苗葉綠素含量，三種白化苗的葉綠素含量趨近於０。綠竹三種白化突變株其31個葉綠體基因經PCR檢測後其重覆數並無減少的現象。但其中6個基因表現量下降。將綠竹三個突變種31個PCR產物進行定序分析，三種個白化苗現有的葉綠葉綠體基因組的核苷酸序列並沒有突變發生。利用生物晶片大範圍觀察基因表現差異，綠竹三種白化苗分別篩選到表現明顯下降的基因。目前了解綠竹白化苗發生原因不同於其他組織培養系統，但尚未能了解其機制。

Building the system of bamboo tissue culture reproduction could propagate and provide for virus-free bamboo plantlet. But propagating bamboo plantlet in tissue culyure, there were producing albinisms in subculture. Cause of many species albino mutant in tissue culture regenerating from single-cell microspore culture was chloroplast genome deletion. But the cause of bamboo albinism through nodal bub culture prorpagating was not published. The object of this study was capitalized bamboo somatic cell mutants and using polymerase chain reaction, reverse transcription polymerase chain reaction and microarray technology to investigate albinisms. psaB (PSI P700 apoprotein A2) DNA was 17% compare with wild type among the 8 genes that copy numbers decreased in in Dendrocalamus Iatiflorus Munro albino mutant. There was no gene copies number decreased of 31 chloroplast genomic genes in Bambusa oldhamii albinism but 6 genes expression has gone down. In addition, using microarray has screened genes expression decreased obviously in Bambusa oldhamii three albinism mutamt. The cause of albinisms were difference between Bambusa oldhamii and other tissue culture system, but the mechanism was unclear in present.