Scott Gillian M, Barrell Barclay G, Oram Jon, Rawlinson William D
Virology Division, Department of Microbiology SEALS, Prince of Wales Hospital, Randwick NSW, Australia.
Virus Genes. 2002;24(1):39-48. doi: 10.1023/a:1014033920070.
The genome of human cytomegalovirus (HCMV) has been studied extensively in some regions, but not others. In this study, transcripts of the genome were further characterised for open reading frames (ORFs) TRL7, UL36, UL65, UL94, US3 and US34, and for the previously unrecognised ORF, UL20a. Reverse transcription-PCR demonstrated the presence of spliced transcripts from the putative glycoprotein gene, UL20a, at early and late times post-infection. US3 full-length and spliced transcripts, including a previously unidentified transcript (US3ii), were described at immediate early times. Sequencing of the complete ORFs of UL20a and US3 from 21 clinical isolates showed that US3 is well conserved in all isolates (97-100% identity), whereas UL20a shows more variation at the nucleotide level, with 90-100% identity. The limits of transcription, and splice donor and acceptor sequences for UL20a and US3 were conserved in all isolates, indicating likely conservation of mRNA splicing patterns. Sequencing a late cDNA library identified the limits of transcription for ORFs TRL7, UL94 and US34 and transcription from the TRL7 ORF was confirmed by northern blotting. Transcripts were found that were congruent with UL36 and UL65, but these differed in the limits previously predicted for these ORFs. These findings show the variation between predicted and actual transcription and indicate the complex nature of transcription from HCMV ORFs.
人类巨细胞病毒（HCMV）的基因组在某些区域已得到广泛研究，但在其他区域并非如此。在本研究中，对基因组转录本的开放阅读框（ORF）TRL7、UL36、UL65、UL94、US3和US34，以及之前未被识别的ORF UL20a进行了进一步表征。逆转录聚合酶链反应（RT-PCR）证明，在感染后的早期和晚期，来自假定糖蛋白基因UL20a的剪接转录本均存在。在即刻早期发现了US3全长和剪接转录本，包括一个先前未鉴定的转录本（US3ii）。对21株临床分离株的UL20a和US3完整ORF进行测序表明，US3在所有分离株中保守性良好（同一性为97 - 100%），而UL20a在核苷酸水平上变异更大，同一性为90 - 100%。所有分离株中UL20a和US3的转录界限以及剪接供体和受体序列均保守，表明mRNA剪接模式可能保守。对晚期cDNA文库进行测序确定了ORF TRL7、UL94和US34的转录界限，Northern印迹法证实了来自TRL7 ORF的转录。发现了与UL36和UL65一致的转录本，但这些转录本在先前预测的这些ORF的界限上有所不同。这些发现显示了预测转录与实际转录之间的差异，并表明HCMV ORF转录的复杂性。
