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Molecular Biotechnology

, Volume 53, Issue 2, pp 150–158 | Cite as

Molecular Studies on the Transmission of Indian Cassava Mosaic Virus (ICMV) and Sri Lankan Cassava Mosaic Virus (SLCMV) in Cassava by Bemisia tabaci and Cloning of ICMV and SLCMV Replicase Gene from Cassava

  • Raghu Duraisamy
  • Senthil NatesanEmail author
  • Raveendran Muthurajan
  • Karthikayan Gandhi
  • Pugalendhi Lakshmanan
  • Nageswari Karuppusamy
  • Mohan Chokkappan
Research

Abstract

Cassava mosaic disease, caused by cassava mosaic geminiviruses are transmitted by Bemisia tabaci. The B. tabaci adults from colonies reared on virus free cassava plant produced from apical meristem culture was studied to determine their ability to transmit Indian cassava mosaic virus (ICMV) and Sri Lankan cassava mosaic virus (SLCMV) from cassava to cassava. Virus free plants were confirmed by polymerase chain reaction (PCR) using geminivirus degenerate primers. The virus acquisition access period (AAP) of 48 h on virus infected cassava leaves and 48 h virus inoculation access periods on virus free healthy leaves were investigated. Both ICMV and SLCMV were absolutely transmitted by whiteflies reared on cassava. Virus specific primers were designed in the replicase region and used to detect virus in B. tabaci after different AAP. The PCR amplified replicase genes from virus transmitted cassava leaves were cloned the plasmid DNA was isolated from a recombinant colony of E. coli DH5α after their confirmation by colony PCR and sequenced them. The nucleotide sequences obtained from automated DNA sequencing were confirmed as ICMV and SLCMV replicase gene after homology searching by BLAST and found to be a new isolates. The nucleotide sequences of new isolates were submitted in GenBank (accession number JN652126 and JN595785).

Keywords

B. tabaci Cloning Meristem culture Sequencing Virus transmission 

Abbreviations

AAP

Acquisition access period

ACMD

African cassava mosaic disease

ACMV

African cassava mosaic virus

BAP

N-6-Benzylaminopurine

Bp

Base pair

BLAST

Basic local alignment search tool

Cm

Centimetre

CMD

Cassava mosaic disease

CMGs

Cassava mosaic geminiviruses

CMV

Cassava mosaic virus

DNA

Deoxy ribonucleic acid

EACMCV

East African cassava mosaic Cameroon virus

EACMKV

East African cassava mosaic Kenya virus

EACMMV

East African cassava mosaic Malawi virus

EACMV

East African cassava mosaic virus

EACMZV

East African cassava mosaic Zanzibar virus

EtBr

Ethidium bromide

GA3

Gibberellic acid

IAP

Inoculation access period

ICMV

Indian cassava mosaic virus

M

Molar

μg

Microgram

ml

Millilitre

μl

Microliter

μM

Micromolar

mm

Millimeter

NCBI

National center for biotechnology information

ng

Nanogram

SACMV

South African cassava mosaic virus

SDS

Sodium dodecyl sulfate

SLCMV

Sri Lankan cassava mosaic virus

MS

Murashige and Skoog

MCM

Meristem culture medium

NAA

1-naphthalene acetic acid

PCR

Polymerase chain reaction

RH

Relative humidity

TNAU

Tamil Nadu Agricultural University

Notes

Acknowledgments

The authors would like to thank Rajiv Gandhi National Fellowship (RGNF) grant (No: F. 14-2 (SC)/2008 (SA-III) Dt: 14.09.2009) to Mr. D. Raghu from the University Grant Commission (UGC), New Delhi, India for financial support. We express our sincere thanks to Dr. K. Shanmugam, Professor and Head for providing CMD infected cassava stakes maintained at Tapioca and Castor Research Station (TCRS), Yethapur at Salem in Tamil Nadu. We are also grateful to the Professor and head, Department of Plant Molecular Biology and Biotechnology (DPMBB), Centre for Plant Molecular Biology (CPMB), Tamil Nadu Agricultural University (TNAU) for providing the laboratory facilities.

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Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  • Raghu Duraisamy
    • 1
  • Senthil Natesan
    • 1
    Email author
  • Raveendran Muthurajan
    • 1
  • Karthikayan Gandhi
    • 2
  • Pugalendhi Lakshmanan
    • 3
  • Nageswari Karuppusamy
    • 4
  • Mohan Chokkappan
    • 5
  1. 1.Department of Plant Biotechnology, Centre for Plant Molecular Biology and BiotechnologyTamil Nadu Agricultural UniversityCoimbatoreIndia
  2. 2.Centre for Plant Protection StudiesTamil Nadu Agricultural UniversityCoimbatoreIndia
  3. 3.Horticulture College and Research InstituteTamil Nadu Agricultural UniversityCoimbatoreIndia
  4. 4.Tapioca and Castor Research StationTamil Nadu Agricultural UniversitySalemIndia
  5. 5.Central Tuber Crops Research InstituteTrivandrumIndia

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