Abstract
BackgroundThe genome of Banana bunchy top virus (BBTV) consists of at least six circular, single-stranded DNA components of ~ 1 kb in length. Some BBTV isolates may also carry satellite DNA molecules that are not essential for BBTV infection. The relation between multipartite DNA virus replication and their transcriptional levels and the underlying mechanism remain unclear.ResultsTo understand the coordinated replication and transcription of the multiple genomic components, the absolute amounts of each BBTV DNA component were measured by real-time PCR (qPCR), and their transcriptional levels were determined by RNAseq and reverse transcription-qPCR (qRT-PCR). Significant differences were found in the absolute amounts of individual BBTV genomic components. Transcriptional levels of each BBTV genomic component obtained from the RNAseq data matched closely to those obtained from qRT-PCR, but did not correspond to the absolute amount of each DNA component. The ratio of transcript over DNA copies ranged from 46.21 to 1059.44%, which was possibly regulated by the promoter region in the intergenic region of each component. To further determine this speculation, the promoter region of the DNA-S, −M or -N was constructed to the upstream of green fluorescent protein (GFP) gene for transient expression by agrobacterium-mediated transformation method. The qRT-PCR showed the highest transcriptional activity was promoted by DNA-N promoter, about 386.58% activity comparing with CaMV 35S promoter. Confocal microscopy observation showed that the intensity of green fluorescence was corresponding to that of qRT-PCR.ConclusionsOur data clearly showed that BBTV was able to control the transcriptional level of each DNA component independently by through the promoter sequences in the intergenic region. Moreover, a cis-acting element from DNA-N component had a high transcriptional activity.
Highlights
The genome of Banana bunchy top virus (BBTV) consists of at least six circular, single-stranded DNA components of ~ 1 kb in length
59.28, 51.46, 56.13 and 56.86 million raw reads were generated from B2, B4, H4 and H5 banana leaves respectively, and 56.22, 48.75, 53.34 and 52.61 million clean reads were further generated of these samples
According to the value of RPKM calculated from the mapping data, DNA-N (852,540.40), DNA-S (631,875.55) and DNA-U3 (564,461.46) components were highly expressed, followed by the DNA-M (111,881.29) with moderate expression
Summary
The genome of Banana bunchy top virus (BBTV) consists of at least six circular, single-stranded DNA components of ~ 1 kb in length. Some BBTV isolates may carry satellite DNA molecules that are not essential for BBTV infection. Its genome contains at least six circular single-stranded DNA components of ~ 1 kb in size [4,5,6,7,8,9,10]. Some BBTV isolates may carry 1 ~ 3 satellite DNA components that are not essential for BBTV infectivity [11, 12]. With the exception of DNA-U3, each BBTV genomic component encodes a single protein with known functions. The satellite molecules encode an assistant replication initiation protein (RepA) with a function similar to that of the Rep [11, 12]
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