本文關鍵詞： 大黃 ISSR SRAP 體系優(yōu)化 親緣關系分析　出處：《河南科技學院》2017年碩士論文　論文類型：學位論文

【摘要】：菜用大黃(Rheum rhaponticum L.)屬于多年生草本植物,其葉柄粗壯多汁,富含纖維素A、維生素C、鈣、磷以及琥珀酸等營養(yǎng)物質(zhì),是理想的保健蔬菜,在歐洲、北美和大洋洲地區(qū)大面積用于商業(yè)栽培。十七世紀,中國藥用大黃被引進至歐洲后經(jīng)過長期的自然進化演變?yōu)椴擞么簏S,然而關于菜用大黃與藥用大黃親緣關系未見報道。本研究對22份大黃品種(6份菜用大黃和16份藥用大黃)進行了種子形態(tài)學觀測和ISSR及SRAP分子標記分析。通過菜用大黃ISSR-PCR反應體系的優(yōu)化及驗證、ISSR及SRAP標記引物的篩選、親緣關系分析等研究,對菜用大黃和藥用大黃的親緣關系進行了研究,并初步揭示菜用大黃的起源,為課題組今后的育種工作提供重要的理論依據(jù)。主要研究結(jié)果如下:(1)建立了菜用大黃ISSR-PCR最佳反應體系。即10×PCR Buffer2.5μL,模板DNA125ng,Mg2+2.5mmol/L,dNTPs 0.35mmol/L,引物0.5μmol/L,Taq DNA聚合酶0.04U/μL,反應體系的總體積為25μL。(2)通過對22個大黃品種的種子長度、寬度、翅寬和千粒重四個形態(tài)指標進行測量,利用SAS v9.2軟件進行聚類分析。結(jié)果顯示當最大距離為3.5時,可以將22份材料劃分為4類,其中RT品種的種子在四個指標上明顯與其他品種存在較大的差異。(3)利用菜用大黃品種R34和藥用大黃品種SC分別對100條ISSR引物和182對SRAP引物組合進行篩選,共獲得多態(tài)性好,譜帶清晰,結(jié)果重復性高的ISSR引物13條、SRAP引物組合32對。ISSR-PCR擴增共獲得74個等位基因位點,其中59個是多態(tài)性位點;SRAP-PCR擴增共計獲得227個條帶,其中206條具有多態(tài)性。結(jié)果表明這兩種分子標記均能提供豐富的遺傳信息,均可用于大黃遺傳多樣性分析。(4)利用POPGENE1.32軟件對22份大黃品種進行遺傳相似性和遺傳距離分析,ISSR-PCR分子標記得出大黃各品種間的遺傳相似系數(shù)在0.3220-0.8983之間,遺傳距離在0.1072-0.9865之間;其中,SC和YN遺傳相似性系數(shù)最大,說明這兩者之間的親緣關系最近;土大黃和RG的相似性系數(shù)最小,表明這二者之間的親緣關系最遠。SRAP-PCR分子標記得出大黃品種間的遺傳相似性系數(shù)在0.3301-0.8495之間;其中,RG和RO遺傳相似性系數(shù)最大,說明這兩者之間的親緣關系最近;南大黃和RO的相似性系數(shù)最小,表明這二者之間的親緣關系最遠。(5)利用NTSYS pc2.10c軟件對22份大黃品種的親緣關系進行聚類分析,基于ISSR及SRAP兩種標記分析數(shù)據(jù)綜合聚類結(jié)果,在相似性系數(shù)0.60處可將22份材料分成4類。第一類為R54、R34和RO,均為菜用大黃;第二類包括川大黃、苦大黃、毛大黃、R19、SC、ZJ、HN、GZ和HB,除R19外均為藥用大黃;第三類為南大黃、西寧大黃、山大黃、馬蹄大黃、RG、RT、HU、SX和YN,其中,RG和RT為菜用大黃;土大黃單獨聚在了第四類。
[Abstract]:Rheum rhaponticum L.) is a perennial herb with a thick, juicy petiole, rich in nutrients such as cellulose A, vitamin C, calcium, phosphorus and succinic acid. It is an ideal health vegetable in Europe. In 17th century, Chinese medicinal rhubarb was introduced to Europe and evolved into vegetable rhubarb after a long period of natural evolution. However, the relationship between vegetable rhubarb and medicinal rhubarb was not reported. In this study, the seed morphology and ISSR and SRAP molecular markers of 22 rhubarb varieties (6 vegetable rhubarb and 16 medicinal rhubarb) were observed. The ISSR-PCR reaction system of rhubarb and the screening of SRAP marker primers were optimized and verified. The relationship between vegetable rhubarb and medicinal rhubarb was studied, and the origin of vegetable rhubarb was preliminarily revealed. The main results are as follows: 1) the optimal reaction system of Rhubarb ISSR-PCR for vegetable is established, that is, 10 脳 PCR Buffer2.5 渭 L, mg 22.5 mmol / L mg 22.5 mmol / L, primer 0.5 渭 mol / L Taq DNA polymerase 0.04 U / 渭 L, primer 0.5 渭 mol / L Taq DNA polymerase 0.04 U / 渭 L. The seed length of 22 rhubarb varieties was determined by 25 渭 L. The width, wing width and 1000-grain weight were measured and cluster analysis was carried out with SAS v9.2 software. The results showed that 22 samples could be classified into 4 groups when the maximum distance was 3.5. The seeds of RT varieties were obviously different from other varieties in four indexes.) 100 ISSR primers and 182 pairs of SRAP primer combinations were screened by using R34 and SC, respectively, and a good polymorphism was obtained. The results showed that there were 74 alleles in 32 pairs of ISSR primer combinations with high reproducibility, 59 of which were polymorphic loci and 227 were amplified by SRAP-PCR. Of these, 206 were polymorphic. The results showed that both of these markers could provide abundant genetic information. It can be used to analyze the genetic diversity of Rhubarb. 4) the genetic similarity and genetic distance of 22 Rhubarb cultivars were analyzed by POPGENE1.32 software. The genetic similarity coefficient was 0.3220-0.8983 and the genetic distance was 0.1072-0.9865. The genetic similarity coefficient of SC and YN is the largest, which indicates that the genetic relationship between them is the closest, and the similarity coefficient between Rhubarb and RG is the smallest. The genetic similarity coefficient was 0.3301-0.8495, and the genetic similarity coefficient between RG and RO was the largest, which indicated that the genetic relationship between RG and RO was the most recent, and the genetic similarity coefficient of RG and RO between the two cultivars was the most distant, which indicated that the genetic similarity coefficient was between 0.3301-0.8495 and 0.3301-0.8495, respectively. The similarity coefficient between South rhubarb and RO was the smallest, indicating that the genetic relationship between the two cultivars was furthest. The genetic relationship of 22 Rhubarb cultivars was analyzed by NTSYS pc2.10c software. The clustering results were synthesized based on the data of ISSR and SRAP markers. At the similarity coefficient of 0.60, 22 samples can be classified into four categories. The first type is R54, R34 and RO. they are both vegetable rhubarb, the second are Radix et Rhizoma et Rhizoma Chuan, Radix et Rhizoma Kuihuang, Radix et Rhizoma Rhei, R19SCC, JJ, HNN, GZ and HBB, all of which are medicinal rhubarb except R19, the third category is Nanda Huang and Xining Rhubarb. Radix rhubarb, Rhizoma et Rhizoma horseshoe, RGG, RGG, HUU, SX and YN, among them, RG and RT are used as vegetable rhubarb, and Rhubarb is clustered in 4th classes alone.
【學位授予單位】：河南科技學院
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：S644.9;S567.239

【參考文獻】

相關期刊論文 前10條

1 張德純;;食用大黃[J];中國蔬菜;2017年02期

2 王洪玲;梁文娟;鐘國躍;梁健;;小大黃根部酚類成分的研究[J];中成藥;2016年10期

3 李永平;康建坂;林琿;;番茄ISSR-PCR反應體系的優(yōu)化與驗證[J];農(nóng)學學報;2016年03期

4 徐世榮;潘鶴立;吳嘉玲;丁安琪;呂恃衡;潘東明;余磊;張智偉;李小婷;;‘紅肉蜜柚’果實汁胞粒化過程中POD同工酶譜分析[J];果樹學報;2016年04期

5 楊金東;甘玉偉;;甘南州唐古特大黃資源現(xiàn)狀、問題及保護措施[J];甘肅科技縱橫;2015年06期

6 李宗艷;秦艷玲;蒙進芳;唐岱;王錦;;西南牡丹品種起源的ISSR研究[J];中國農(nóng)業(yè)科學;2015年05期

7 昝逢剛;應雄美;吳才文;趙培方;陳學寬;馬麗;蘇火生;劉家勇;;98份甘蔗種質(zhì)資源遺傳多樣性的AFLP分析[J];中國農(nóng)業(yè)科學;2015年05期

8 田艷伶;李志輝;楊模華;張斌;;鉤栗ISSR-PCR反應體系的建立與優(yōu)化[J];中南林業(yè)科技大學學報;2015年02期

9 徐世松;王呈丹;黃素榮;張志群;黃少華;楊光穗;;紅掌種質(zhì)資源形態(tài)學標記與遺傳多樣性分析[J];熱帶作物學報;2014年10期

10 李瑞峰;高鵬;朱子成;欒非時;;基于形態(tài)學標記及SSR標記的甜瓜主栽品種分類鑒定研究[J];中國蔬菜;2014年06期

，

本文編號：1529801