Naslov
The pilot study of viroids in asymptomatic Solenostemon scutellarioides (L.) Codd plants

Autori
Škorić, Dijana ; Černi, Silvija ; Jezernik, Karlo

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Izvornik
The Interplay of Biomolecules HDBMB2014, Congress of the Croatian Society of Biochemistry and Molecular Biology, Book of Abstracts / Katalinić, Maja ; Kovarik, Zrinka - Zagreb : Hrvatsko Društvo za Biotehnologiju, 2014, 130-130

ISBN
978-953-95551-5-1

Skup
The Interplay of Biomolecules HDBMB2014, Congress of the Croatian Society of Biochemistry and Molecular Biology

Mjesto i datum
Zadar, Hrvatska, 24.09.2014. - 27.09.2014

Vrsta sudjelovanja
Poster

Vrsta recenzije
Domaća recenzija

Ključne riječi
Coleus blumei viroids; RT-PCR; cloning; SSCP; sequencing

Sažetak
Solenostemon scutellarioides (L.) Codd is a new name of the ornamental plant Coleus blumei Benth., well known worldwide for its brightly coloured leaves and medicinal value owing to its phenolic content. It is also a main host of six Coleus blumei viroids (CbVds) species (CBVd-1 to -6). CbVd-1 occurs globally. CbVd-2 and -4 have been reported only in Germany, CbVd-3 in Germany and China, and CbVd-5 and -6 only in Southeast Asia. Interestingly, CbVds are transmissible not only by vegetative propagation and mechanically like other viroids, but also by seeds. Viroids are noncoding small covalently closed circular RNA (cccRNA) molecules independently parasitizing in plants. Most of the viroids, including CbVds, belong to the family Pospiviroidae having over 50% internal base pairing in their cccRNAs, Central Conserved Region (CCR), asymmetric rolling–circle replication catalysed by cellular RNA polymerase II in nucleus and no ribozyme activity. The aim of this study was to investigate the occurrence of Pospiviroidae in a small population of asymptomatic S. scutellarioides plants grown in the Botanical garden of the Faculty of Science, Zagreb. Leaf and stem tissue from ten plants were used to compare three methods for obtaining viroid RNAs. Generic primers covering CCR of all six CbVds were used in RT-PCR based detection procedure. Although the method using microliter quantities of crude sap can be useful for preliminary detection, and the total RNA after RNeasy kit (Qiagen) extraction was acceptable too, the best results were obtained from small RNA templates after procedure encompassing phenol-chloroform extraction, LiCl-fractionation and CF-11 cellulose chromatography. Amplicons of approx. 250 and 360 bp were obtained from 5, and 2, out of ten plants, respectively. As direct sequencing of PCR products did not yield clear results due to the quasispecies nature of viroids, an amplicon of each size was AT-cloned. After SSCP-screening, all identified variants in each viroid’s population structure were sequenced. For the first time in Croatia, CbVd-1 and CbVd-3 were identified with variants of 249-251 and 361-362 nucleotides, respectively. The incidence levels of 50% for CbVd-1 and 20% for CbVd-3 are high for such a small population of investigated plants but those values are within reported incidence ranges. Also, finding CbVd-3 the second time in Europe and in 2 out of only 10 plants suggests CbVd-3 could be more widespread than assumed.

Izvorni jezik
Engleski

Znanstvena područja
Biologija

Napomena
Dio istraživanja proveden je u sklopu izrade diplomskog rada studenta diplomskog studija molekularne biologije Karla Jezernika.

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