【摘要】：柑橘是世界上重要的水果,柑橘的無核及晚熟性狀一直以來都是柑橘育種工作者重點(diǎn)關(guān)注內(nèi)容。最近本課題組在福建省發(fā)現(xiàn)一個(gè)名為‘拋橘’的無核柑橘地方資源,其在當(dāng)?shù)鼐哂薪倌甑姆N植歷史,但文獻(xiàn)未有記載�！畳侀佟哂泄馑瞥�,果皮有褶皺且易剝離,果實(shí)大小與葡萄柚相類似的特點(diǎn),果實(shí)成熟期一般在次年一月中旬左右,可掛果至次年4月中旬,單果均重349.8g,可食率平均為45.9%,可溶性固形物含量為10.4%,可滴定酸含量為1.62%。綜上,‘拋橘’具有較高的研究和開發(fā)價(jià)值。本課題研究以‘拋橘’為研究對(duì)象,從雌雄配子體育性等方面初步研究了‘拋橘’無核的原因;利用形態(tài)學(xué)觀察和SSR標(biāo)記對(duì)‘拋橘’進(jìn)行遺傳鑒定。取得的主要研究結(jié)果如下:一、‘拋橘’的雌雄配子體發(fā)育正常1.連續(xù)兩年,觀察花粉育性指數(shù)。使用醋酸洋紅染色觀察‘拋橘’的花粉染色活力,結(jié)果顯示正常率均在80%以上;利用培養(yǎng)基上培養(yǎng)‘拋橘’花粉,結(jié)果顯示萌芽率均在75%以上;利用血球計(jì)數(shù)板統(tǒng)計(jì)其每花藥花粉量為2×104粒左右。2.解剖學(xué)結(jié)果表明:‘拋橘’雄配子體發(fā)育正常。掃描電鏡結(jié)果顯示‘拋橘’花粉形態(tài)正常,并且正常率可達(dá)到80%以上。3.石蠟切片觀察:觀察了‘拋橘’雌配子體的四個(gè)發(fā)育時(shí)期,未發(fā)現(xiàn)敗育及畸形發(fā)育的情況;對(duì)‘拋橘’雄配子體的發(fā)育進(jìn)行了觀察,其發(fā)育正常。綜上,‘拋橘’的無核原因可能為自交不親和或胚胎敗育。二、‘拋橘’的遺傳鑒定1.7對(duì)cp SSR引物和13對(duì)nSSR引物分別對(duì)24和22個(gè)柑橘屬及其近緣屬植物進(jìn)行了UPGMA聚類分析,推測(cè):‘拋橘’是以柚類(Citrus maxima)×酸橙類(citrus aurantium)雜交而來。2.基因重測(cè)序數(shù)據(jù)分析顯示:?基因重測(cè)序數(shù)據(jù)分析顯示:‘拋橘’與酸橙的基因組序列相似度在85%以上,推測(cè)‘拋橘’可能屬于酸橙的一種;盡管葉片形態(tài)、果實(shí)外形以及果肉質(zhì)地和色澤與溫州蜜柑相近,但與溫州蜜柑基因組序列相似度只有63.49%,不能發(fā)現(xiàn)存在直接親緣關(guān)系的證據(jù)。3.13對(duì)nSSR引物對(duì)38個(gè)柑橘屬及其近緣屬植物進(jìn)行了UPGMA聚類分析,結(jié)果顯示:‘拋橘’和代代酸橙有較密切的親緣關(guān)系。推測(cè):代代酸橙可能是‘拋橘’的一種類型或者是雜交親本之一。三、q PCR檢測(cè)4個(gè)與SI相關(guān)基因在‘拋橘’不同組織中的表達(dá)量結(jié)果顯示:Cg Rns6、Cg Rns9、Cg Rns L在‘拋橘’不同組織中的表達(dá)沒有規(guī)律,而Cg RNS3在‘拋橘’的雌蕊中特異性表達(dá),表明‘拋橘’無核的原因可能是自交不親和,而Cg RNS3可能是其自交不親和的候選基因。
[Abstract]:Citrus is an important fruit in the world. The seedless and late maturing traits of citrus have always been the focus of citrus breeders. Recently, our research group discovered a seedless citrus local resource called 'throwing orange' in Fujian Province, which has a planting history of nearly a hundred years, but it has not been recorded in the literature. 'throwing orange' has the characteristics of orange flesh, fold and easy peeling, and the fruit size is similar to that of grapefruit. The fruit ripening period is generally in the middle of January of the following year. The average weight of single fruit is 349.8g, the average edible rate is 45.9%, the soluble solid content is 10.4%, and the titratable acid content is 1.62%. In summary, 'throwing orange' has a high research and development value. In this study, the reason of seedlessness of 'throwing orange' was studied from the aspects of male and female gametes, and the genetic identification of 'throwing orange' was carried out by morphological observation and SSR markers. The main results are as follows: first, the male and female gametophyte development of 'throwing orange' is normal. The pollen fertility index was observed for two years in a row. The pollen staining activity of 'throwing orange' was observed by magnitine acetate staining, the results showed that the normal rate was above 80%, the germination rate of 'throwing orange' pollen was above 75%, and the amount of anther powder per anther was about 2 脳 104 grains by blood cell counting board. The anatomical results showed that the male gametophyte developed normally. The results of scanning electron microscope (SEM) showed that the pollen morphology of 'throwing orange' was normal, and the normal rate could reach more than 80%. Paraffin section observation: the four developmental stages of female gametophyte of 'throwing orange' were observed, and no abortion and abnormal development were found, and the development of male gametophyte of 'throwing orange' was observed, and the development of male gametophyte was normal. In summary, the seedless cause of orange throwing may be self-incompatibility or embryo abortion. 2. UPGMA cluster analysis of 24 and 22 citrus and their related plants was carried out with 1.7 pairs of cp SSR primers and 13 pairs of nSSR primers, respectively. it was inferred that 'throwing orange' was made by (citrus aurantium) hybrid of grapefruit (Citrus maxima) 脳 lime. 2. The analysis of gene resequencing data shows that:? The analysis of gene resequencing data showed that the similarity of genome sequence between 'throwing orange' and lime was more than 85%. It was speculated that 'throwing orange' might belong to a kind of lime. Although the leaf morphology, fruit shape, pulp texture and color were similar to those of Satsuma mandarin, the similarity of genome sequence with Satsuma mandarin was only 63.49%, and no evidence of direct genetic relationship could be found. 3.13 pairs of nSSR primers were used to analyze 38 citrus and their related plants by UPGMA cluster analysis. The results showed that there was a close genetic relationship between 'throwing orange' and generation lime. It is speculated that the generation lime may be a type of 'throwing orange' or one of the hybrid parents. The expression of four SI related genes in different tissues of 'throwing orange' was detected by three, q PCR. The results showed that the expression of Cg Rns6,Cg Rns9,Cg Rns L in different tissues of 'throwing orange' was irregular, while the specific expression of Cg RNS3 in the pistil of 'throwing orange' indicated that the reason for the lack of nucleus in 'throwing orange' may be self-incompatibility, while Cg RNS3 may be a candidate gene for self-incompatibility.
【學(xué)位授予單位】：華中農(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)】：S666

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