Nicotiana plumbaginifolia 與紫外線照射之 N. sylvestris 間的體細胞雜交

Abstract

本實驗以紫外線照射 Nicotiana sylvestris ，原生質體，再與硝酸鹽還原?缺失的 N . plumbaginifolia 原生質體融合，希望能夠得到此二菸草之不對稱體細胞雜種 ( asymmetric somatic hybrids ）。以不同劑量（ 0 、 500 、 1000 、 2000 J/ m2 ）紫外線處理 N . sylvestris 的原生質體，分別與 N . plumbaginifolia 原生質體融合後，以硝酸鹽為唯一氮源的 MC 培養基作篩選，且劑量在 500 J / m2 以上的紫外線可完全抑制 N . sylvestris的細胞分裂，故在紫外線與 MC 培養基的處理下，所得的再生植株應皆為體細胞雜種。且由染色體鑑定與南方氏雜交分析結果皆顯示所得到的再生植株是二親本的體細胞雜種。而紫外線劑量愈高，融合後所得到的癒合組織愈少，體細胞雜種中 N . sylvestris的染色體丟失數目愈多，且染色體構造變異也愈多。但 N . sylvestris的染色體丟失數目還是很有限，可能在紫外線的處理上需有所修改，如降低紫外線之 dose rate ，提高紫外線劑量等 。

Fusion between protoplasts from a nitrate reductase deficient mutant of Nicotiana plumbaginifolia and UV-irradiated protoplasts of N. sylvestris was carried out for the purpose of obtaining asymmetric somatic hybrids. A comparison was made between the effects of UV doses in the range 0 to 2000 J / m2 as a pretreatment of isolated N. sylvestris protoplasts. After fusion, protoplasts were cultured in media containing nitrate as the sole nitrogen source to select the somatic hybrids. Chromosomal identification and Southern hybridization analysis revealed that all regenerated plants were somatic hybrids. The results also revealed that a dose of 500 J/m2 UV could prevent further division of N. sylvestris protoplasts. Besides, an increase in UV irradiation doses applied prior to fusion led to a decrease in green calli formation. Furthermore, the loss of N. sylvestris chromosomes and the occurrence of chromosomal structural changes in somatic hybrids were directly correlated with UV doses. However, the amounts of the N. sylvestris genome lost were limited. The reason for this is discussed.