臺灣金線連遺傳變異之簡單序列重複及限制酶片段長度多型性分析   全文下載

Genetic Variation of Anoectochilus formosanus Revealed by ISSR and AFLP Analysis

台灣金線連為台灣民間昂貴草藥，經常被用於高血壓、糖尿病及肝炎的治療。本研究依據其外表形態特徵，收集紅莖、綠莖、長葉和圓葉四個金線連品系，進行簡單序列重複標誌的篩選。同時發現，其中以長葉品系的多醣體含量最高（3.39 ± 0.74%，多醣體克重/植物乾克重），而紅莖品系的多醣體含量最低（1.27 ± 0.41%），相差達兩倍以上。為了瞭解金線連組織培養的體細胞遺傳變異性，挑選民間栽培常見的長葉品系，以同一母本的20株分生苗，進行限制?片段長度多型性分析。本研究的目的欲篩選多型性的標誌，作為將來培養高多醣體含量之台灣金線連的育種工具。本研究的結果顯示，81條簡單重複序列引物的其中6條引物，共計產生26個多型性標誌，足以有效區分該四個品系。另一方面，在限制?片段長度多型性分析中，利用17組引物偵測金線連分生苗的體細胞遺傳變異，計算得到這20株分生苗的變異介於0~5.43%之間；其中以B6引物組偵測金線連體細胞變異的能力最高，而E4引物組為最低。這些具有偵測力的aFlP引物組，將是篩選台灣金線連組織培養之最低體細胞變異分生苗的有利工具。

Anoectochilus formosanus is an expensive medicinal folk herb in Taiwan. The plant has been used to treat hypertension, diabetes, and hepatitis by native people for a long time. In this study, 4 typical lines of A. formosanus, red-stem, green-stem, narrow-leaf, and round-leaf, were collected and screened to identify the line-specific SSR (single sequence repeat) markers. In addition, we found the narrow-leaf line exhibited the highest polysaccharide content (3.39 ± 0.74%, polysaccharide wt/plant dried wt) which is more than twofold of the lowest content of the red-stem line (1.27 ± 0.41%). In the culture program construction, the extent of the somaclonal variation among 20 regenerated shoots from the same parental individual of a narrow-leaf line was revealed using the amplified fragment length polymorphism (AFLP) technique. The aim of this study is to preliminarily investigate the genetic variation of A. formosanus through polymorphic marker screening, which will benefit to the cultivation of A. formosanus with high polysaccharide content in future work. The results showed that 6 out of 81 primers screened, generating a total of 26 markers, were able to differentiate the 4 lines in the inter single sequence repeat (ISSR) analysis. On the other hand, 17 sets of AFLP primers were chosen to detect the somatic variation among the excised shoots derived from the same individual. The genetic variation among the regenerated shoots ranged from 0.00% to 5.43%. The primer pair B6 exerts the most powerful selection ability in the AFLP analysis among 20 somaclonal samples used, whereas the E4 primer pair the lowest. The AFLP primer pairs become useful tools for selecting the lowest somaclonal variation from the regenerated shoots of A. formosanus under the tissue culture program.