Studies on the Physiological Function of Lymphocytes and the Polymorphism of Major Histocompatibility Complex in Japanese Quails

1.对日本鸣禽淋巴细胞抗原的研究：(1)以5个胸腺细胞克隆QT-1-QT-5和5个法氏囊细胞克隆QB-1-QB-5为杂交体。(2)以抗胸腺细胞克隆QB-1-QB-5为免疫器官进行免疫组织化学染色。QT-1^+、-4^+和5^+细胞仅存在于胸腺皮质，QB-1^+、-2^+和-3^+细胞存在于法氏囊、延髓和脾PARS的B细胞部位。QT-1^+、-4^+和5^+细胞仅存在于胸腺，其出现频率为70-80%。2.日本鹌鹑主要组织相容性复合体(MHC)I类基因的分析：(1)以BF10为探针，以鸡MHC I类基因为探针，对日本鹌鹑基因组DNA进行RFLPs分析，得到9个阳性克隆：QF41、63、76、95、103、111、113、128，以BF10和QF41为随机序列，筛选出与日本鹌鹑MHC I类基因对应的9个阳性克隆。(3)以基因组DNA为模板，以QF63和QF41为模板，对QF63和QF41的扩增产物进行分析，发现QF63和QF41的扩增产物片段大小相差609bp，而QF41扩增产物的扩增片段大小相差434bp。QF63在α1和α2区之间有181bp的内含子，而QF41在α1和α2区之间没有内含子。(4)用两种限制性内切酶BAN II和EcoT14对QF63的扩增产物进行限制性内切酶酶切，得到9种单倍型QF63产物，但只有5种单倍型。

1.Studies on the lymphocyte antigens of Japanese quail : (1) As hybridomer of 5 clones derived from thymocyte : QT-1-QT-5 ; and 5 clones derived from bursacyte : QB-1-QB-5.(2) Imminohistochemical analysis on the immuno organ were carried out by the monoclonal anitibodies. QT-1^+, -4^+ and 5^+ cells only exist incortex of thymus.QB-1^+, -2^+ and -3^+ cells exist in medulla or bursa of Fabricius and in B cell locus of PALS of spleen.(3) Lymphocyte antigens were analyzed by flowcytometory. QT-1^+, -4^+ and 5^+ cells only exist thymus and their frequencies were 70 - 80%. The frequencies of QB-1^+, -2^+ and -3^+ cells were 90% in brusacytes.2.Analysis of major histocompatibility complex (MHC) class I gene from Japanese quail : (1) RFLPs patten of genomic DNA from Japanese quail which were analyzed using BF10 in chicken MHC class I cDNA as a probe showed polymorphism.(2) Nine positive clones : QF41,63,76,95,103,111,113,128 ; corresponding to the MHC class I gene of Japanese quail were isolated by screening of liver cDNA library with BF10 and QF41 as arobe. Based on comparison among sequences of QF clones, MHC class I of Japanese quail classified into 5 loci.(3) PCR analysis of alpha1-alpha2 region using genomic DNA as a template with QF63 and QF41 specific primers were revealed different sizes of fragment by 609bp in PCR product of QF63 vs 434bp in QF41 product. QF63 has 181bp intron between alpha1 and alpha2 regions, but QF41 have not intron between alpha1 and alpha2 regions.(4) PCR products using the QF63 specific primer were digested with the 2 kinds of restriction endnuclease : Ban II and EcoT14 I.Combination of PCR-RFLP patterns using 2 kinds of restriction endnuclease classified 9 haplotypes QF63 product, but only 5 haplotypes ovserved in this experiment.