六種豆象的線粒體Co1基因的檢測及鑒定
發(fā)布時(shí)間：2019-01-10 15:01
【摘要】：四紋豆象與花生豆象、菜豆象、鷹嘴豆象、綠豆象、咖啡豆象形態(tài)特征相似,菜豆象、四紋豆象、鷹嘴豆象、花生豆象、咖啡豆象已被國家列為禁止進(jìn)境的檢疫性有害生物,綠豆象是中俄、中南植檢植保雙邊協(xié)定中規(guī)定的檢疫性害蟲,六種豆象的經(jīng)濟(jì)意義均十分重要,口岸檢驗(yàn)檢疫部門主要以成蟲的外部形態(tài)特征作為主要鑒定依據(jù)。另外,口岸截獲的不單是成蟲,經(jīng)常還有些是卵、幼蟲、蛹等未成熟蟲態(tài),尋找一種正確、快速的鑒定方法在口岸檢驗(yàn)檢疫中的應(yīng)用具有十分重要的指導(dǎo)意義。本研究對象為四紋豆象,首先提取四紋豆象的線粒體基因,通過設(shè)計(jì)引物,對四紋豆象等六種豆象的線粒體特異性基因Co1進(jìn)行提取,測序,通過構(gòu)建系統(tǒng)發(fā)育樹,完成檢疫性豆象類的分子鑒定,然后進(jìn)一步對四紋豆象進(jìn)行線粒體基因組全序列的測定與分析。將有利于建立更穩(wěn)健的系統(tǒng)發(fā)育關(guān)系。實(shí)驗(yàn)結(jié)果如下:(1)試驗(yàn)方法的優(yōu)化:由于常規(guī)檢驗(yàn)豆象的方法不能夠準(zhǔn)確快速地鑒定出豆象的種類的限制,對臨沂出入境檢驗(yàn)檢疫局截獲的豆象以及泰安糧食市場沉積儲(chǔ)藏物(豆類)中的豆象,用PCR級(jí)柱式動(dòng)物線粒體DNAout試劑盒處理,對其線粒體DNA中特異性基因Co1進(jìn)行提取鑒定測序。(2)Co1基因全長420bp,Blast對比分析六種豆象Co1基因的差異。(3)以六種實(shí)驗(yàn)材料豆象特異性基因Co1序列為基礎(chǔ),結(jié)合GenBank上查找的不同豆象的特異性Co1序列構(gòu)建系統(tǒng)發(fā)育樹,加以分析,了解不同豆象的親緣差異,嘗試構(gòu)建參照體系,方便對截獲的豆象及幼蟲卵等通過分子測序確定其所屬種。(4)驗(yàn)證:通過截獲檢疫性豆象卵若成蟲等個(gè)體進(jìn)行分子水平的鑒定及測序代入系統(tǒng)發(fā)育樹,經(jīng)比對確認(rèn)所屬種,完成測序。
[Abstract]:The four-striped bean elephant is similar to the peanut bean elephant, the chickpea elephant, the mung bean elephant, the coffee bean elephant. The bean elephant, the four-striped bean elephant, the chickpea, the peanut bean elephant and the coffee bean elephant have been listed as quarantine pests banned by the state. The economic significance of the six species of legume is very important. The port inspection and quarantine departments mainly take the external morphological characteristics of adults as the main basis for identification. In addition, not only adults but also eggs, larvae, pupae and other immature insects are intercepted at ports. It is of great significance to find a correct and rapid identification method in port inspection and quarantine. Firstly, the mitochondrial gene of tetracarpus was extracted. The mitochondrial specific gene (Co1) was extracted from six species of legume by designing primers, and sequenced, and the phylogenetic tree was constructed. The molecular identification of quarantine legumes was completed, and then the complete mitochondrial genome sequence was determined and analyzed. It will help to establish a more stable phylogenetic relationship. The experimental results are as follows: (1) the optimization of the test method: because of the limitation of the conventional method to identify the species of the elephant accurately and quickly, The beans intercepted by the Entry-Exit Inspection and Quarantine Bureau of Linyi and the legumes in the depositional storage products (legumes) of the Tai'an grain market were treated with PCR grade column animal mitochondrial DNAout kit. The specific gene Co1 in mitochondrial DNA was extracted and sequenced. (2) the total length of Co1 gene was 420 BP blast. (3) based on the Co1 sequence of six experimental materials, the difference of Co1 gene of six soybean elephants was compared. The phylogenetic tree was constructed by combining the specific Co1 sequences of different bean elephants found on GenBank. The phylogenetic differences of different legumes were analyzed and the reference system was constructed. It is convenient to identify the species of the intercepted bean elephants and larval eggs by molecular sequencing. (4) Verification: through the molecular level identification and sequencing of individuals such as the quarantine elephant eggs if the adult worm, and so on, the phylogenetic tree can be identified by comparison and identification of the species to which they belong. Complete the sequencing.
【學(xué)位授予單位】：山東農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：S433.5
本文編號(hào)：2406455
[Abstract]:The four-striped bean elephant is similar to the peanut bean elephant, the chickpea elephant, the mung bean elephant, the coffee bean elephant. The bean elephant, the four-striped bean elephant, the chickpea, the peanut bean elephant and the coffee bean elephant have been listed as quarantine pests banned by the state. The economic significance of the six species of legume is very important. The port inspection and quarantine departments mainly take the external morphological characteristics of adults as the main basis for identification. In addition, not only adults but also eggs, larvae, pupae and other immature insects are intercepted at ports. It is of great significance to find a correct and rapid identification method in port inspection and quarantine. Firstly, the mitochondrial gene of tetracarpus was extracted. The mitochondrial specific gene (Co1) was extracted from six species of legume by designing primers, and sequenced, and the phylogenetic tree was constructed. The molecular identification of quarantine legumes was completed, and then the complete mitochondrial genome sequence was determined and analyzed. It will help to establish a more stable phylogenetic relationship. The experimental results are as follows: (1) the optimization of the test method: because of the limitation of the conventional method to identify the species of the elephant accurately and quickly, The beans intercepted by the Entry-Exit Inspection and Quarantine Bureau of Linyi and the legumes in the depositional storage products (legumes) of the Tai'an grain market were treated with PCR grade column animal mitochondrial DNAout kit. The specific gene Co1 in mitochondrial DNA was extracted and sequenced. (2) the total length of Co1 gene was 420 BP blast. (3) based on the Co1 sequence of six experimental materials, the difference of Co1 gene of six soybean elephants was compared. The phylogenetic tree was constructed by combining the specific Co1 sequences of different bean elephants found on GenBank. The phylogenetic differences of different legumes were analyzed and the reference system was constructed. It is convenient to identify the species of the intercepted bean elephants and larval eggs by molecular sequencing. (4) Verification: through the molecular level identification and sequencing of individuals such as the quarantine elephant eggs if the adult worm, and so on, the phylogenetic tree can be identified by comparison and identification of the species to which they belong. Complete the sequencing.
【學(xué)位授予單位】：山東農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：S433.5
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