Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/22947
標題: 新式生技策略應用於促進雞隻免疫反應及增強傳染性華氏囊病疫苗保護之研究
Enhancing Immunity and Potentiating Infectious Bursal Disease Vaccination by Novel Biotechnology Approaches
作者: 洪哲明
Hung, Che-Ming
關鍵字: Gingyo-san
銀翹散
lactoferrin
infectious bursal disease
transgenic chicken
乳鐵蛋白
傳染性華氏囊病
基因轉殖雞
出版社: 生命科學系所
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摘要: 本試驗目的在探討以”銀翹散”(Gingyo-san, GGS;試驗一)及基因重組乳鐵蛋白(recombinant porcine lactoferrin, rPLF;試驗二)分別在飼糧中添加0.5%, 1% 及2% (w/w)第三種不同之劑量,對免疫過傳染性華氏囊病(infectious bursal disease, IBD)活毒疫苗之雞隻,在週邊淋巴細胞增生與IBD抗體力價之影響;試驗三旨在應用類微脂體(lipofetin)與外源DNA結合所生之複合物,並經由精子載體法或將lentiviral vector注入公雞睪丸方法,嘗試進行產製抗病力強之基因轉殖雞。 試驗一結果顯示低劑量的GGS可以顯著提高IBD抗體力價;而中與高劑量GGS可促進IL-2及INF-γ之產生。此外,試驗亦證明GGS可以改變T淋巴細胞在IL-4與IL-12之表現,從而促使CD4+ T淋巴細胞朝向Th1型發展。 試驗二結果顯示,餵飼PLF高劑量顯著增加血清中之IgA, IgG, 及 IBD 抗體力價 (P<0.05),且不管在PLF高劑量或低劑量處理組均可增進雞隻淋巴細胞之 IFN-γ 及IL-12之表現,因此GGS及PLF分別可增進細胞免疫(cell-mediated immunity),並協助IBD疫苗功能及後續抗病毒之能力。 試驗三在精子載體法部分雛雞在5週齡取其基因體DNA,並經PCR初步證明,35.8%雛雞係攜帶有該外源DNA片段者,不過此等攜有外源DNA雞隻在15週齡時,再採其雞血DNA重覆PCR及南方氏吸漬法分析其結果則不再攜外源DNA。而慢病毒載體(lentiviral vector)注入公雞睪丸方面,經飼養至48月齡時利用活體影像系統觀察睪丸帶有外源EGFP者比率為42% (8/19)。故推測新式的睪丸內注射lentiviral vector感染法,在睪丸之生精作用過程中,外源性基因較易穩定嵌入於生殖細胞之基因組(genome)中。
The purpose of experiment is to investigate the effects of diets supplemented with 0.5%, 1% and 2% of Gingyo-san (GGS) (Exp. 1) and recombinant porcine lactoferrin (rPLF) (Exp. 2) to the peripheral lymphocyte proliferation and IBD antibody titer of IBD vaccinated chicken. In Exp. 3, combination of and Lipofetin and exogenous DNA were applied and tried to produce transgenic chickens via sperm vector method or injection of Lentiviral vector to cock testis. The results in Exp. 1 indicated that low dose GGS significantly increased IBD antibody titer and middle and high dose GGS promoted production of IL-2 and INF-γ. GGS changed the T lymphocyte expression on IL-4 and IL-12 which induced the development of CD4+ T lymphocyte toward Th1 type. The results in Exp. 2 indicated that high dose PLF significantly increased serum IgA, IgG and IBD antibody titer (P<0.05). In addition, both high and low dose PLF increased the expression of IFN-γ and IL-12 of chicken lymphocyte. Hence, GGS and PLF can promote cell-mediated immunity, IBD vaccine function and subsequent antivirus response. In Exp. 3 of sperm vector method, 5-week-old chicks genomic DNA was extracted for PCR test. 35.8% chicks obtained exogenous DNA fragment. Blood of those chickens with exogenous DNA at 15 weeks of age. The results using PCR and southern blotting method showed no more exogenous DNA. In the injection of Lentiviral vector into cock testis, the ratio of testis with exogenous EGFP was 42% (8/19) using live image system. We expected that the new method of testis injecting with Lentiviral vector is available. The foregin DNA is more stably maintained in germ cells during spermatogertein.
URI: http://hdl.handle.net/11455/22947
其他識別: U0005-2101201016291100
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