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應用噬菌體呈現技術於豬隻黃體激素之檢測 = Porcine progesterone detection through phage display technology

紀錄類型:	書目-語言資料,印刷品 : 單行本
並列題名:	Porcine progesterone detection through phage display technology
作者:	李旻宸,
其他團體作者:	國立高雄大學
出版地:	[高雄市]
出版者:	撰者;
出版年:	2012[民101]
面頁冊數:	106面圖，表格 : 30公分;
標題:	噬菌體呈現技術
標題:	phage display technology
電子資源:	http://handle.ncl.edu.tw/11296/ndltd/08036652084244446188
附註:	106年10月31日公開
附註:	參考書目：面74-86
附註:	含附錄
摘要註:	如何使豬隻繁殖達最大效率是提高豬場經濟的主要重點。研究顯示，辨別豬隻懷孕與否的決定因素之一為血液中黃體激素濃度。此荷爾蒙在胚胎順利著床後方穩定的上升，以維持適合胚胎成長的環境直到分娩，與未懷孕母豬受雌激素影響而再度發情導致黃體激素下降有顯著的差異。因此本研究以此做為診斷的指標，配合噬菌體呈現技術分析噬菌體與黃體激素的親和效果，並評估以此取代黃體激素抗體做為檢測試劑成分的潛力。將官能基修飾的黃體激素(progesterone-3-O-CMO)接合至牛血清白蛋白(BSA)後，由西方點墨法結果得知在68.4 kDa左右有顯著的呈色，證實成功將黃體激素接至BSA上。以ph.D.-C7C和ph.D.-7噬菌體胜肽庫對progesterone-3-O-CMO-BSA conjugate進行生物親和性篩選並去除會親和BSA的噬菌體後，篩選範圍縮小至315 pfu與175 pfu；隨機挑選多個噬菌體經擴大、純化後藉酵素聯結免疫吸附法(ELISA)與點墨法(dot blot)分析噬菌體親和progesterone程度，挑出具辨別性之噬菌體定序分析，將胺基酸排列後可得三種較一致性之序列。此外，比對NCBI資料庫中已知的黃體激素受體發現可對應到豬隻黃體激素受體(NP_001159960)與豬隻黃體激素受體膜蛋白(NP_999076)。以純的黃體激素溶液測試此等噬菌體，發現ph.D.-C7C編號74與ph.D.-7編號19等噬菌體對黃體激素分子較具專一性，且其序列可對應到黃體激素受體結合位，因此具有開發鑑別豬隻懷孕檢驗試劑的潛力。 Raising the efficiency of pig-breeding is a major point to increase the economy of pig farm. Several studies showed that the concentration of progesterone in blood was a crucial factor used to determine the pregnancy or not in sow. After embryo implantation, the concentration of this hormone stably rises and keeps in a high level to maintain a suitable environment for embryo development until delivery. It is significantly different with the non-pregnant sow that influenced by estrogen and returned to estrous leading to the decrease of progesterone concentration. Therefore, progesterone was used as a diagnostic marker, and analyzed it with phage display technology to evaluate its potential of replacing the antibody against progesterone and facilitating the development of diagnostic kit. Western blot analysis showed that the molecular weight of progesterone-3-O-CMO-BSA conjugate was 68.4 kDa. It indicated that progesterone was conjugated onto BSA successfully. After 2 rounds of biopanning with ph.D.-C7C and ph.D.-7 phage display peptide libraries and removing the phages bind to BSA, the residual numbers of phage were 315 pfu and 175 pfu, respectively. Several phages were randomly selected, amplified, purified and then assayed by enzyme linked immunosorbent assay (ELISA) and dot blot analysis to evaluate their binding affinity to progesterone. Several phages with high binding affinity were selected for DNA sequences analyses and deduced into amino acid sequences. Three consensus sequences were found after sequences alignment. Moreover, several selected sequences were found corresponding to the progesterone receptor (PGR) of porcine (NP_001159960) and progesterone receptor membrane component 1 (PGRMC1) of porcine (NP_999076) deposited in NCBI database. These phages were assayed with pure progesterone, it showed that phage C7C-74 and D7-19 have higher affinity and specificity to progesterone and their sequences were corresponding to the ligand biding site of porcine PGR. In conclusion, the selected phages have the potential for the development of diagnostic kit in porcine pregnancy detection.

應用噬菌體呈現技術於豬隻黃體激素之檢測 = Porcine progesterone detection through phage display technology
李, 旻宸

應用噬菌體呈現技術於豬隻黃體激素之檢測 = Porcine progesterone detection through phage display technology / 李旻宸撰 - [高雄市] : 撰者, 2012[民101]. - 106面 ; 圖，表格 ; 30公分.
106年10月31日公開參考書目：面74-86含附錄.
噬菌體呈現技術phage display technology

應用噬菌體呈現技術於豬隻黃體激素之檢測 = Porcine progesterone detection through phage display technology
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