Sequencing, Improved Detection, and a Novel Form of Kalanchoë top-spotting virus.
Sequencing, Improved Detection, and a Novel Form of Kalanchoë top-spotting virus.
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DOI：
10.1094/pd-89-0298
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发表时间：
2005-03
期刊：
影响因子：
4.5
通讯作者：
Zihong Yang;M. Nicolaisen;N. Olszewski;B. Lockhart
中科院分区：
文献类型：
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作者：
Zihong Yang;M. Nicolaisen;N. Olszewski;B. Lockhart
采用一种防止病毒在初始提取步骤中丢失的方法，从Kalanchoë blossfeldiana感染的叶片组织中纯化出Kalanchoë顶斑病毒（KTSV）病毒粒子。克隆了病毒双链DNA基因组并对其进行了测序。KTSV基因组大小为7591 bp，包含三个开放阅读框，分别编码21,14和223 kDa的蛋白质。KTSV基因组的大小和组织与其他粉蚧传播的坏病毒相似。基于KTSV基因组序列的几个寡核苷酸引物对被用来有效地检测植物中的病毒，从而消除了可靠筛选kalanchoë繁殖种群和KTSV感染育种系的主要限制。通过聚合酶链反应（PCR）扩增和限制性内切酶酶切得到的扩增子，鉴定和区分了两个KTSV序列，一个是诱导症状的，另一个是不诱导症状的。从移植物传播试验和PCR索引的初步结果表明，无症状形式的KTSV可能代表一个综合的病毒元素。这种整合的副逆转录病毒元件的出现给育种和繁殖种群的常规PCR索引带来了实际问题，也增加了由这些病毒整合物引起的症状性偶发感染的可能性。
Virions of Kalanchoë top-spotting virus (KTSV) were purified from infected leaf tissue of Kalanchoë blossfeldiana using a procedure that prevented loss of virus in the initial extraction step. The double-stranded DNA viral genome was cloned and sequenced. The KTSV genome was 7,591 bp in size and contained three open reading frames capable of encoding proteins of 21, 14, and 223 kDa, respectively. The size and organization of the KTSV genome were similar to those of other mealybug-transmitted badnaviruses. Several oligonucleotide primer pairs, based on the KTSV genomic sequence, were used to efficiently detect the virus in plants, thereby removing a major constraint to reliable screening of kalanchoë propagating stock and breeding lines for KTSV infection. Two KTSV sequences, one symptom-inducing and the other not, were identified and differentiated by polymerase chain reaction (PCR) amplification and digestion of the resulting amplicon with restriction endonucleases. Preliminary results from graft-transmission tests and PCR indexing suggest that the nonsymptomatic form of KTSV may represent an integrated viral element. The occurrence of such integrated pararetroviral elements poses practical problems for routine PCR indexing of breeding and propagating stock, and also raises the possibility of symptomatic episomal infections arising from these viral integrants.