为揭示牛MHC-DRB3上游调控区多态对其表达调控作用,利用PCR-SSCP和DNA序列测定技术分析并建立BoLA-DRB3*exon2和BoLA-DRB3 URR之间的连锁关系。结合不同近端调控区与转录因子结合模式预测结果,选择3种不同上游调控序列的连锁组合,应用实时定量RT-PCR技术对牛MHC-DRB3基因mRNA相对表达量进行了验证分析。结果表明,上游调控区基因型为H17H17的BoLA-DRB3基因mRNA相对表达量显著高于(P<0.01)基因型H1H1及H5H6。初步证明上游调控区中的多态会引起结构基因表达量的变化。
王学清
,
李魁英
,
裴翠娟
,
张峰
,
吴占军
,
刘小虎
,
马书林
,
张新同
,
王昆
. 中国荷斯坦牛BoLA-DRB3基因的mRNA表达研究[J]. 华北农学报, 2013
, 28(4)
: 19
-22
.
DOI: 10.3969/j.issn.1000-7091.2013.04.004
To study the regulation of upstream regulatory region polymorphisms on BoLA- DRB3 gene expres- sion，PCR- SSCP and DNA sequencing were used to identify the linkage relationship between BoLA- DRB3* exon2 and BoLA- DRB3 URR．Combined with the predicted results of binding mode between different URR and transcrip- tion factor，three linkage types were selected to detect the BoLA-DRB3 mRNA levels by relative Real- time PCR ex- periments． The results showed that the expression abundances of BoLA-DRB3 varied in different linkage types.The expression level of mRNA of BoLA- DRB3 gene linked with H17H17 was significant higher than H1H1 and H5H6 (P＜0.01)．This preliminary proved that the polymorphism of upstream regulatory region can cause the expression change of BoLA- DRB3 structural gene．
[1] Klein J． Origin of MHC Polymorphisms: the trans-species hypothesis[J]. Human Immunology，1987，19: 155-62．
[2] Babbit B P，Allen P M． The binding of immunogenetic peptides to Ia histocompatibility molecules[J]. Nature，1987，317: 359-362．
[3] Buus S，Sette A． The relation between major histocompati-bility complex (MHC) restriction and the capacity of Ia to bind immunogenetic peptides[J]. Science，1987，235: 1353-1355．
[4] Schmittgen T D，Zakrajsek B A． Effect of experimental treatment on housekeeping gene expression: validation by real-time，quantitative RT-PCR[J]. J Biochem Biophys Methods，2000，46(1-2): 69-81．
[5] Russell G C． BoLA class nucleotide sequences，1996: report of the ISAG BoLA Nomenclature Committee[J]. Animal Genetics，1997，28: 169-180．
[6] Anim Gelhaus，Sequence and PCR-RFLP analysis of 14 novel BoLA-DRB3 alleles[J]. Animal Genetics，1995，26: 147-153．
[7] Wang Kun，Sun Dong xiao，Zhang Yuan． Sequencing of 15 new BoLA-DRB3 alleles[J]. International J Immunogenet-ics，2008: 331-332．
[8] 王学清，王 昆，王沛宇，等． 中国荷斯坦牛MHC-DRB3 * exon2 的 PCR-SSCP 多态性分析[J]. 华北农学报，2010，25(5): 94-98．
[9] 王 昆，孙东晓，徐如海，等． BoLA-DQB 上游调控区多 态性研究[J]. 畜牧兽医学报，2005，36(6): 521-525．
[10] Wang K，Sun D X，Li K Y，et al． Identification of four novel alleles of the BoLA-DRB3 upstream regulatory re-gion in Chinese yellow cattle[J]. Tissue Antigens，2012，80: 58-60．
[11] Ripoli M V，Peral-Garcia P，Dulout F N，et al． Polymor-phism in the bovine BoLA-DRB3 upstream regulatory re-gions detected through PCR-SSCP and DNA sequencing [J]. Gene，2004，339: 71-78．
