人类及动物RNA病毒的反向遗传系统

反向遗传系统可以对RNA病毒直接进行遗传操作,为RNA病毒的分子生物学研究提供了一种强大的工具。在过去20年,特别是自90年代中期第一例负链RNA病毒感染性克隆构建成功以来,动物RNA病毒的分子生物学研究取得了长足的进展,这很大程度上归功于各种动物RNA病毒反向遗传系统的建立。这里系统总结了人类及动物非反转录RNA病毒中各类代表性成员在建立反向遗传系统时的方案设计、遇到的困难及研究者如何克服这些困难。分类讨论到的代表性病毒种属有脊髓灰质炎病毒、冠状病毒（包括SARS病毒）、黄病毒、野田村病毒、流感病毒、传染性法氏囊病病毒以及呼肠孤病毒等。     

Conserved Region of Mammalian Retrovirus RNA

The viral RNAs of various mammalian retroviruses contain highly conserved sequences close to their 3' ends. This was demonstrated by interviral molecular hybridization between fractionated viral complementary DNA (cDNA) and RNA. cDNA near the 3' end (cDNA(3')) from a rat virus (RPL strain) was fractionated by size and mixed with mouse virus RNA (Rauscher leukemia virus). No hybridization occurred with total cDNA (cDNA(total)), in agreement with previous results, but a cross-reacting sequence was found with the fractionated cDNA(3'). The sequences between 50 to 400 nucleotides from the 3' terminus of heteropolymeric RNA were most hybridizable. The rat viral cDNA(3') hybridized with mouse virus RNA more extensively than with RNA of remotely related retroviruses. The related viral sequence of the rodent viruses (mouse and rat) showed as much divergence in heteroduplex thermal denaturation profiles as did the unique sequence DNA of these two rodents. This suggests that over a period of time, rodent viruses have preserved a sequence with changes correlated to phylogenetic distance of hosts. The cross-reacting sequence of replication-competent retroviruses was conserved even in the genome of the replication-defective sarcoma virus and was also located in these genomes near the 3' end of 30S RNA. A fraction of RD114 cDNA(3'), corresponding to the conserved region, cross-hybridized extensively with RNA of a baboon endogenous virus (M7). Fractions of similar size prepared from cDNA(3') of MPMV, a primate type D virus, hybridized with M7 RNA to a lesser extent. Hybridization was not observed between Mason-Pfizer monkey virus and M7 if total cDNA's were incubated with viral RNAs. The degree of cross-reaction of the shared sequence appeared to be influenced by viral ancestral relatedness and host cell phylogenetic relationships. Thus, the strikingly high extent of cross-reaction at the conserved region between rodent viruses and simian sarcoma virus and between baboon virus and RD114 virus may reflect ancestral relatedness of the viruses. Slight cross-reaction at the site between type B and C viruses of rodents (mouse mammary tumor virus and RPL virus, 58-2T) or type C and D viruses of primates (M7, RD114, and Mason-Pfizer monkey virus) may have arisen at the conserved region through a mechanism that depends more on the phylogenetic relatedness of the host cells than on the viral type or origin. Determining the sequence of the conserved region may help elucidate this mechanism. The conserved sequences in retroviruses described here may be an important functional unit for the life cycle of many retroviruses.