应用mRNA差异显示技术寻找调控白腰纹鸟前脑发声控制神经核RA性双态性发育的基因(英文)

Sexually dimorphic expression of the genes in the forebrain vocal-control nucleus RA of juvenile white-rumped munia Lonchura striata by mRNA differential display technique

鸣禽的发声行为及其前脑发声控制核团存在显著的性双态性 ,这种性双态性是在发育过程中逐步建立起来的。鸣禽白腰纹鸟 35日龄 (P35 )前 ,雌、雄鸟前脑发声控制神经核RA (Robustnucleusofarcopallium)不存在性别差异 ;到 4 5日龄 (P4 5 )时 ,雌、雄鸟RA体积已经出现了显著的差异。推测这种神经结构的性双态性的形成与各种基因的不同时空表达有关。本实验以发育不同阶段 (P35时的雌性 ,P4 5时的雌性和雄性 )的雌、雄白腰纹鸟RA核团为研究对象 ,用mRNA差异显示银染技术来寻找前脑上述三组材料间的差异表达基因 ,试图发现调控RA核团性双态性发育的关键基因。本实验共选用了 4种锚定引物与 16种随机引物所组成的 6 4组引物进行DDRT PCR (mRNAdifferentialdisplayandreversetranscription polymerasechainreaction)反应。第一次扩增后 ,在测序胶上显示出 115 8条cDNA扩增产物 ,其中 2 73个为差异表达。长度分布于 10 0 - 5 0 0bp之间的差异条带共 16 1条。本实验重点研究了P35雌性和P4 5雄性共表达或P4 5雌性单独表达的条带 (共 19条 ) ,推测这些基因片段可能与引起RA核团中的细胞凋亡和性双态性的形成有关。二次PCR扩增后 ,进一步筛选出 16个差异条带 ,其中 11个为P35雌性和P4 5雄性共同表达的 ,5个是P4 5雌?

Song behavior and its song-control nuclei (such as the robust nucleus of the arcopallium-RA) are highly sexually dimorphic in passerine songbirds. These dimorphisms are not present early in development, but arise through processes during sensitive periods in life. There are no sex differences in RA volume and neuron numbers of white-rumped munia at P35 (date after birth), but sexual dimorphism in RA volume became very markedly at P45. It is thought that the development of neural dimorphism is controlled by genes. In this experiment, the female and male RAs at developmental ages P35 (females only) and P45 (females and males) were studied by mRNA differential display (DD) technology. For the DDRT-PCR, combinations of the four T_ 12MN primers together with sixteen arbitrary primers, AP1-16, were used. This analysis yielded 1 158 bands in total, of which 273 bands were differentially generated. There were 161 differential bands which size was between 100 bp and 500 bp. The aim of this study was focused on the cDNAs co-expressed in P35 female and P45 male (sixty-one bands) or expressed only in female at P45 (nineteen bands), which might play a role in the dimorphic development of RA. After the second PCR amplification, sixteen differential bands were screened out, of which eleven were co-expressed in P35 female and P45 male, five were only expressed in female at P45, which might underlie the genetic mechanism of the RA dimorphic development in white-rumped munia. A differential segment named T3, which had 81% homology with Bcl complex in chichen, might be a member of Bcl-2 protein family or relate to the retaining of RA volume, was chosen to sequence and analyse. However, the information about T3 segment in detail needed further research. These results show that mRNA DD methodology can represent a potential tool for investigation of the gene expression in the brain of songbird development .