草蝦基因組的遺傳分析:種源鑑定及遺傳圖譜之建立

Genetic analysis of tiger shrimp genome: stock identification and construction of genetic linkage map

尤恩民(En-Min You)

指導教授 ： 于宏燦

國立臺灣大學/生命科學院/動物學研究所/博士(2008年)

https://doi.org/10.6342/NTU.2008.01304

摘要

草蝦(Penaeus monodon)是全球養殖產業中最重要的海產物種之一，也是具有生物多樣性研究價值的海洋生物。因此，本研究利用分子遺傳標記去研究草蝦基因組的特性，包括兩個大主題: (1)草蝦地理族群的遺傳多樣性與種源分析。(2)草蝦遺傳圖譜的建構。
本論文共分為四章，第一章包括研究主題之文獻回顧，與研究方法的論述，並論文的研究方向。論文的第二章，為利用分子遺傳標記進行野生草蝦族群遺傳多樣性的研究成果，此部分已發表於Animal Genetics期刊中。論文的第三章則為草蝦遺傳圖譜建構之研究成果。論文的第四章則為本研究的總結論與未來的展望。
論文的第一個主題-草蝦地理族群遺傳與種源分析的研究(論文第二章)，是使用來自於8個不同地理區域，一共355隻野生草蝦作為實驗樣本，以自行開發的10個微隨體標記及粒線體控制區(mtCR)序列，來分析草蝦族群與個體的遺傳分化情形。結果顯示，不論是使用微隨體標記或粒線體控制區(mtCR)序列進行地理親緣關係分析，均得到相同的結論，即西印度洋(馬達加斯加及肯亞)與西太平洋(台灣、越南、菲律賓、澳洲及泰國東岸泰國灣)的草蝦族群在遺傳上已有明顯分隔。此項研究結果對於草蝦養殖產業的種源鑑定應大有助益。
論文的第二個主題-草蝦遺傳圖譜的建構(論文第三章)，是利用多型性微隨體標記，輔以多型性AFLP標記，以一個isofemale F1家族(內含280個子代個體)為樣本，共使用148個草蝦微隨體標記以及134個AFLP標記，進行基因型分析及連鎖分析，建構出草蝦雌性及雄性遺傳圖譜。雌性遺傳圖譜含128個微隨體標記及55個AFLP標記，並組成了46個連鎖群，而連鎖群的總遺傳長度是2248.9 cM；雄性遺傳圖譜含126個微隨體標記及79個AFLP標記，並組成了43個連鎖群，而連鎖群的總遺傳長度是2286.8 cM。這兩個遺傳圖譜共同擁有106個標記，並座落於36個同源連鎖群(homologous linkage group)。此遺傳圖譜將能應用於數量性狀基因座(quantitative trait loci; QTL]的定位與分子育種選殖。

關鍵字

草蝦 ； 微隨體 ； 粒線體控制區序列 ； 遺傳多樣性 ； 遺傳圖譜 ； AFLP ； isofemale F1家族 ； 同源連鎖群

並列摘要

The black tiger shrimp (Penaeus monodon) is an ecologically and economically important marine crustacean and is widely distributed in the Indo-Pacific region. Here I investigated the genome of P. monodon in two aspects, i.e., (1) the genetic diversity of wild populations, (2) the construction of a genetic linkage map.
There are 4 chapters in this thesis. Chapter 1 contained the paper review of the research topics, the research methods, and the research aspects. In chapter 2 the genetic diversity and the stock identification of wild P. monodon populations in Indo-Pacific region was revealed, and this part of work has been published in Animal Genetics. In chapter 3 the constructed linkage maps of the tiger shrimp based on microsatellites and AFLP markers were presented. Chapter 4, the final chapter, was concerned with the research summary and future perspectives.
In order to investigate the genetic diversity of wild P. monodon populations, tiger shrimps (n = 355) from eight geographical regions were genotyped using of 10 microsatellite markers. A high level of genetic variability (average HO: 0.638-0.743) and significant genetic differentiation (Pairwise FST and RST values) among the populations were revealed. The non-metric multidimensional scaling analysis indicated the presence of three geographic groups in the Indo-Pacific region, i.e., the African populations, a population from western Thailand, and the remaining populations as a whole. In addition, the phylogenetic analysis using the sequences from the mitochondrial control region (mtCR) revealed that the West Indian Ocean populations were genetically differentiated from the West Pacific populations, consistent with the result obtained from the microsatellite data.
In order to acquire more genetic information of tiger shrimp genome, the linkage maps of male and female tiger shrimp (P. monodon) were constructed based on microsatellite and amplified fragment length polymorphism (AFLP) markers. 621 microsatellite and 1151 AFLP markers were tested for polymorphism. Consequently, 372 (167 microsatellite and 203 AFLP markers) were used for genotyping an isofemale F1 family comprising two parents and 280 progeny. From the linkage analysis (LOD score= 4.5), a male and a female linkage map consisting of 43 and 46 linkage groups were constructed, respectively. The male map consisted of 126 microsatellite markers and 79 AFLP markers, whereas the female map consisted of 128 microsatellite markers and 55 AFLP markers. These two maps shared 36 homologous linkage groups.

並列關鍵字

Penaeus monodon ； microsatellite ； mitochondrial control region ； genetic diversity ； linkage map ； amplified fragment length polymorphism (AFLP) ； isofemale F1 family ； homologous linkage groups

參考文獻

Alcivar-Warren A.M.-M., D. , Park S.W., Xu Z., Delaney M. & Zuniga G. (2007) ShrimpMap: A low-density, microsatellite-based linkage map of the pacific whiteleg shrimp, Litopenaeus vannamei: Identification of sex-linked markers in linkage group 4. Journal of shellfish research 26, 1259-77.

Avise J.C. (1994) Molecular Markers, Natural History and Evolution. Chapman & Hall, New York, U.S.A.

Avise J.C. (2004) Molecular Markers, Natural History, and Evolution. Sinauer Associates, Inc., Sunderland, U.S.A.

Benzie J.A.H. (1998) Penaeid genetics and biotechnology. Aquaculture 164, 23-47.

Benzie J.A.H. (2000) Population genetic stucture of Penaeid prawns. Aquaculture Research 31, 95-119.