本計畫使用先前執行的畜禽功能性基因體與蛋白質體計畫成果，並利用近年發展之基因體技術研究早期胚基因表現差異，探討胚細胞基因表現之調控機制，獲得之成果則有利於開發母豬繁殖性能的蛋白質檢測套組及具差異表現之基因型檢測套組，冀能實際與快速地縮短種豬選育時程，並藉此提升豬隻繁殖效能。針對母豬配種前血清中蛋白質形成2D電泳膠片，利用去年已產製之B2M (Beta-2-Microglobulin)單株抗體進行western blot之先導實驗，根據先導實驗結果發現母豬配種前血清中B2M出現八個不同PI位點(不同的isoforms)，其中兩個位點在高產母豬血清中清楚辨認，在低產母豬血清中則難以辨認，我們繼續執行盲樣測試，發現此一檢測方式可以在離乳母豬血清中有效預測下一胎次的高低產情形，未來可以開發成為母豬配種前的健康檢查套組。我們另外也在種豬場收集種母豬繁殖性能資料，選出高繁與低繁性能母豬群，在配種前取其血樣中的白血球，於配種後七天取其血樣中的白血球及blastocyst胚胎後製備RNA，進行44K晶片檢測其基因表現差異，並予以分析獲取高繁與低繁性能表現相關的差異表現基因群。本計畫依據這些資料進行多個基因的啟動子區域及轉錄調控區的預測，找出該區段的單一核?酸多型性(SNP)位點，針對這些位點進行引子組設計，然後進行大量基因型鑑定，使用最佳線性無偏估測(BLUP)估計出母豬繁殖性能的育種價，進行各位點基因型與育種價的相關性分析，完成一組母豬繁殖性能蛋白質測試檢驗套組雛形，以及一組與差異基因regulatory SNP檢測套組一組，以便未來針對母豬進行繁殖性狀的遺傳性能檢測，也能夠提供豬場進行母豬離乳後懷孕健康狀況進行檢測，以提高豬場內種母豬的整體繁殖性能。 In this project, the results from previous studies of?functional genomics and proteomics in livestock animals were used to?detect those?differential gene expression at the early stages of porcine embryogenesis?for?studying the regulatory machnism of the gene expression in embryo. The results obtained will profit the development of?proteim testing kits and SNP?genotyping?sets?of differential gene expression for examining sow reproductive performance.?The 2D gel of proteins?in serum sampled?collected from sows?right after weaning (before next mating)?was used to conduct western blot analysis?by the B2M (Beta-2-Microglobulin) monoclonal antibody produced?in this project. Our pilot study showed that there are eight?spots with different PI values for?B2M protein detected in the western blot analysis in which two spots of B2M isoforms were easily to identify high-productive or low-productive sows. Then, a blind test has been conducted to?identify which sows?will?be high-productive or low-productive in the next parity.?The?B2M?testing method effectively?predict the?high or low?litter size in their next parity.?This?testing kit?is expected to improve the sow reproductive efficiency.?We collected the reproductive performance data?to find out those high-productive and low-productive sows. The RNA samples of?white blood cells were collected from the sows before mating.?And, the RNA samples of white blood cells and blastocyst embryos were collected from the same sows?7 days after mating. The 44K?microarray analyses were conducted using those RNA samples to find out those differentially expressed genes.?The promoter region and their?regulatory elements were predicted in those?genes.?Then, those single nucleotide polymorphsim (SNP) were scanned?in those promoter?regions. According those SNP sites, primer sets were designed?for large-scale genotyping thereafter.?The Best Linear Unbiased Prediction (BLUP) was used to estimate the breeding values of litter size for those?sows. An association analysis was conducted to find those influencial SNPs?for sow reproductive performance. The project for this year is expecting to accomplish a preliminary protein testing kit for sow reproductive performance to practically test in the field. In addition, there will be a set of regulatory?SNPs of differentially expressed genes related to reproductive performance of sows established. These two testing sets (protein and SNP) will be used in testing?genetically reproductive success in sows as well as testing sow's?health status before mating to increase the total reproduction performance of sows on-farm.
