抑制脯胺酸合成酵素基因表現或施用根毛農桿菌對菸草銅累積之影響

Abstract

在許多污染土壤皆可見過量銅存在，植物可能因銅毒害而無法生長，也可能透過生物累積進入食物鏈中，進而造成人體之危害。植生復育是利用植物體對於汙染物的吸收進行土壤復育的技術。但植體於過量重金屬逆境下，會進行生理調節機阻抗重金屬進入。研究顯示脯胺酸的增加是其中一種阻抗機制，導致重金屬吸收能力受抑制，亦有研究指出細胞壁是金屬吸附的第一線位置。本研究利用帶有脯胺酸合成基因反股核苷酸片段之菸草鑲嵌病毒 (pTMV-antiP5CS) 降低菸草在乾旱逆境下葉片脯胺酸的累積，以期望達到銅累積增加之效。試驗結果顯示在5 mM硫酸銅48小時處理下，接種pTMV-antiP5CS之菸草植株對銅的吸收量較對照組菸草多。於一公分根尖菸草吸附試驗中觀察到，經pTMV-antiP5CS接種之菸草細胞壁對於4 mg L-1 銅離子處理的吸附能力較對照組菸草差，推測接種pTMV-antiP5CS抑制菸草脯胺酸合成酵素的作用可能會導致根部細胞壁對於銅離子的吸附能力降低，進而使銅離子易於進入植體內。另施用根毛農桿菌於栽培介質中，發現施用OD600 0.01菌液濃度處理的菸草植體，在2 mM硫酸銅48小時處理下，其銅累積量較無菌液處理組高，推測可能為根毛農桿菌可使毛狀根增生之故。總結而言，抑制脯胺酸合成酵素，導致菸草根尖細胞壁對於銅的吸附能力降低。施用根毛農桿菌之菸草，在2 mM硫酸銅處理下，銅累積量增加，可能是毛狀根細胞壁對銅吸附力較低，也可能是根吸收面積增大所致，尚待進一步的試驗釐清。

Many heavy metal contaminated soils contain excess copper in Taiwan. The mechanisms of plant against the toxicity could be used in phytoremediation. Many studies indicated under heavy metals stress, the accumulation of proline in plant could tolerate the toxicity and reduce the heavy metal absorption and the root cell wall was the main location of heavy metal adsorption. The objective of this study was to use virus-induced gene siliencing (VIGS) technique and inoculate Agrobacterium rhizogenes to Nicotiana benthamiana and to see if the N. benthamiana can accumulate copper under high concentration of copper treatment. The results showed the copper concentration of cell wall of 1 cm root tips in inoculating proline synthesis gene (pTMV-antiP5CS) was lower than control under 4 mg L-1 copper treatment. However, the copper concentration of shoot with inoculation pTMV-antiP5CS tobacco was higher than that of control under 5 mM CuSO4 treatment. These results indicated that the cell wall of 1 cm root tips in pTMV-antiP5CS tobacco plant had smaller copper adsorption capability. The shoot of pTMV-antiP5CS tobacco plant accumulated greater copper compared with the wild type. N. benthamiana inoculated with OD600 0.01 A. rhizogenes had higher concentration of copper compared with that of without inoculation under 2 mM CuSO4 solution treating for 48 h. N. benthamiana treated with A. rhizogenes induced the hair root formation and enhanced the adsorption area in plant root.