本文選題：穿心蓮 + 等離子體處理��； 參考：《廣州中醫(yī)藥大學(xué)》2017年碩士論文

【摘要】：等離子體處理種子技術(shù)已廣泛使用于多種作物,被證實(shí)可提高種子活力,促進(jìn)植物生長(zhǎng),提高植物抗脅迫能力。等離子體處理種子的作用目前主要認(rèn)為是通過(guò)刻蝕種皮來(lái)提高種皮通透性與放電過(guò)程產(chǎn)生的帶電粒子對(duì)種子表面的消毒作用,與抗氧化酶活性提高有關(guān),提高了種子的健壯度;在分子層面的探討幾乎空白,需要深層次挖掘分析。穿心蓮來(lái)源于爵床科(Acanthaceae)植物穿心蓮(Andrographis paniculata(Burm.f.)Nees)的地上部分,干燥后為臨床常用中藥材。研究報(bào)道等離子體處理穿心蓮種子可提高的出苗率和整齊度,促進(jìn)植物生長(zhǎng)速率。本研究系統(tǒng)地考察了等離子體處理對(duì)穿心蓮種子萌發(fā)、出苗、生長(zhǎng)速率的影響,從分子層面考察了等離子體處理對(duì)種子萌發(fā)過(guò)程基因表達(dá)的影響,為揭示等離子體處理種子技術(shù)的作用機(jī)理提供理論基礎(chǔ)。本論文的主要研究?jī)?nèi)容和結(jié)果如下:1.等離子體處理促進(jìn)穿心蓮種子萌發(fā)及幼苗生長(zhǎng)采用大氣壓條件下低電壓(30 V～50 V)高頻交變電場(chǎng)激發(fā)常溫空氣等離子體處理穿心蓮種子,考察了激發(fā)電壓、處理時(shí)間、處理后存放時(shí)間等多個(gè)因素,篩選促進(jìn)其萌發(fā)的最佳條件,探究了等離子體處理對(duì)穿心蓮萌發(fā)、出苗、生長(zhǎng)、等多方面的影響。用30 V激發(fā)的等離子體處理種子3 s后放置4～6 d再置種,發(fā)芽勢(shì)顯著提高;放置時(shí)間過(guò)長(zhǎng)再置種,處理的效果下降或消失。等離子體處理穿心蓮種子的效應(yīng)主要表現(xiàn)在發(fā)芽勢(shì)顯著提高、出苗加快、出苗率升高、幼苗及植株生長(zhǎng)量增加、生殖生長(zhǎng)加快及抗逆性增強(qiáng)。2.等離子體處理后穿心蓮種子萌發(fā)過(guò)程中基因表達(dá)概況本論文采用第二代高通量測(cè)序儀Illumina HiSeq 2500進(jìn)行測(cè)序,共得到106.34 Gb 的 Clean Data,84749 條 Unigene,平均長(zhǎng)度為 758.03 bp,注釋到 Nr、KEGG、GO、Swiss-prot、COG等公共數(shù)據(jù)庫(kù)的Unigene數(shù)共36567條,建立了穿心蓮種子轉(zhuǎn)錄組平臺(tái)。對(duì)對(duì)照組和處理組的吸脹期(分別為a,A)、萌動(dòng)期(b,B)、萌發(fā)高峰期(c,C)三個(gè)階段差異基因進(jìn)行stem趨勢(shì)聚類分析,發(fā)現(xiàn)對(duì)照組和處理組顯著富集的趨勢(shì)相同,表明等離子體處理并未改變種子萌發(fā)過(guò)程基因表達(dá)的主要表達(dá)趨勢(shì),整體遵循種子萌發(fā)基因表達(dá)特有的規(guī)律性變化。等離子體處理與未處理穿心蓮種子相對(duì)應(yīng)三個(gè)萌發(fā)過(guò)程時(shí)間點(diǎn)的轉(zhuǎn)錄組比較發(fā)現(xiàn):在萌動(dòng)期,處理組和對(duì)照組有125個(gè)差異表達(dá)基因,主要富集到物質(zhì)代謝相關(guān)KEGG通路;差異表達(dá)基因主要GO富集到翻譯、初級(jí)代謝過(guò)程、有機(jī)物代謝過(guò)程、代謝過(guò)程、氧化還原過(guò)程等,調(diào)控種子萌發(fā)物質(zhì)代謝,與KEGG富集情況一致。分別對(duì)對(duì)照組和處理組中這125個(gè)基因間達(dá)顯著相關(guān)的基因做基因相關(guān)性圖,對(duì)照組和處理組這125個(gè)基因間的相關(guān)性關(guān)系存在巨大差別,體現(xiàn)了等離子體對(duì)基因表達(dá)調(diào)控關(guān)系的影響。本文對(duì)萌發(fā)過(guò)程中處理組與對(duì)照組相同階段特有差異表達(dá)的基因進(jìn)行了篩選,并進(jìn)行富集分析,KEGG富集結(jié)果發(fā)現(xiàn)處理組從吸脹期到萌動(dòng)期富集通路差異較大,處理組植物激素信號(hào)轉(zhuǎn)導(dǎo)富集基因最多;在對(duì)照組則不在前10條富集通路,對(duì)照組中核糖體通路富集到基因數(shù)最多;從萌動(dòng)期到萌發(fā)高峰期,兩組基因富集情況類似,等離子體處理在種子萌發(fā)過(guò)程的調(diào)控主要表現(xiàn)在種子萌發(fā)的前期,即吸脹期到萌動(dòng)期。對(duì)a_b(對(duì)照組從吸脹期到萌動(dòng)期)、b_c(對(duì)照組從吸脹期到萌動(dòng)期)、A_B(處理組從吸脹期到萌動(dòng)期)、B_C(處理組從吸脹期到萌動(dòng)期)這4個(gè)階段特有的差異表達(dá)中物質(zhì)代謝基因與該階段特有的植物激素、轉(zhuǎn)錄因子、病程相關(guān)基因進(jìn)行基因表達(dá)相關(guān)性分析,結(jié)果顯示,對(duì)照組與處理組間基因調(diào)控網(wǎng)絡(luò)具有顯著差異性,等離子體處理改變了種子萌發(fā)過(guò)程中這些關(guān)鍵基因的表達(dá)調(diào)控模式。本實(shí)驗(yàn)選擇了與種子萌發(fā)相關(guān)的6個(gè)基因進(jìn)行qPCR,結(jié)果顯示,這些基因的表達(dá)趨勢(shì)模式與轉(zhuǎn)錄組測(cè)序結(jié)果大部分一致,說(shuō)明測(cè)序可靠性較高。
[Abstract]:Plasma treatment seed technology has been widely used in a variety of crops, which have been proved to improve seed vigor, promote plant growth and improve plant resistance to stress. The main effect of plasma treatment of seeds is to improve the seed surface permeability and discharge through the etching seed skin to improve the disinfection of the seed surface. It is used to improve the activity of antioxidant enzymes and improve the robustness of the seeds; the discussion at the molecular level is almost blank and needs deep mining analysis. The Andrographis paniculata is derived from the upper part of the Acanthaceae (Andrographis paniculata (Burm.f.) Nees). After drying, it is used as a commonly used Chinese medicinal material. The effects of plasma treatment on the seed germination, emergence and growth rate of Andrographis paniculata were systematically investigated. The effects of plasma treatment on the expression of the seed germination process were investigated from the molecular level to reveal the plasma treatment seeds. The main contents and results of this paper are as follows: 1. plasma treatment promotes the seed germination and seedling growth of Andrographis paniculata by using low voltage (30 V ~ 50 V) high frequency alternating electric field under atmospheric pressure to excite the seed of Andrographis paniculata at normal temperature air plasma, and investigate the excitation voltage and treatment time. A number of factors such as storage time after treatment were used to screen the best conditions to promote the germination. The effects of plasma treatment on the germination, emergence and growth of Andrographis paniculata were investigated. The germination potential was significantly higher after 3 s of plasma treated with 30 V, and the germination potential was significantly higher. The effect of plasma treatment on the seed of Andrographis paniculata was mainly manifested in the remarkable improvement of the germination potential, the expedite emergence of the seedlings, the increase of the seedling rate, the increase of seedling and plant growth, the accelerated growth of the reproductive growth and the increase of the resistance to stress in the seed germination of the seed of Andrographis paniculata after.2. plasma treatment. The second generation high throughput sequencing instrument Ill was used in this paper. Umina HiSeq 2500 was sequenced. A total of 106.34 Gb Clean Data, 84749 Unigene, and an average length of 758.03 BP were obtained. A total of 36567 public databases such as Nr, KEGG, GO, Swiss-prot, COG and other public databases were set up. The three phase difference gene of C (C) was analyzed by stem trend cluster analysis. It was found that the trend of significant enrichment in the control group and the treatment group was the same, which showed that the plasma treatment did not change the main expression trend of gene expression in the seed germination process. Compared with the transcriptional group at the time point of three germination process, it was found that there were 125 differentially expressed genes in the treatment group and the control group at the germination stage, which mainly enriched the KEGG pathway related to the material metabolism, and the main GO of the differential expression genes was enriched in the translation, the primary metabolic process, the organic metabolism process, the metabolic process, and the redox process. And so on, the regulation of the metabolism of seed germination material was consistent with the enrichment of KEGG. The correlation between the 125 genes in the control group and the treatment group was significantly related to the genes. The correlation of the 125 genes between the control group and the treatment group had a huge difference, which reflected the influence of the plasma on the regulation of gene expression. The genes of specific differential expression in the same stage of the treatment group and the control group were screened, and the enrichment analysis was carried out. The enrichment results of KEGG enrichment found that the enrichment pathway of the treatment group from the bloating period to the germinating stage was different, and the plant hormone signal transduction was the most abundant in the treatment group; in the control group, the first 10 enrichment pathways and the control groups were not in the control group. The number of genes in the middle nuclear sugar pathway is the most. From the germinating period to the peak period, the two sets of gene enrichment are similar. The regulation of the plasma treatment in the seed germination process is mainly in the early stage of seed germination, that is, the bloating period to the germinating period, and the a_b (the control group from the bloating period to the germinating period), and the b_c (the control group from the bloating period to the germinating period), A_B (treatment group from suction period to germinal period), B_C (treatment group from suction period to germination stage) of the 4 stages of the specific differential expression of material metabolism gene and the specific plant hormones, transcription factors, disease related genes gene expression correlation analysis, the results show that the control group and the treatment group gene regulation network has a significant difference. The expression regulation mode of these key genes during seed germination was changed by plasma treatment. 6 genes related to seed germination were selected for qPCR. The results showed that the trend patterns of these genes were most consistent with that of the transcriptional group, and that the reliability of the sequencing was higher.

【學(xué)位授予單位】：廣州中醫(yī)藥大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：S567.219;Q943.2

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