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ARS Home » Pacific West Area » Parlier, California » San Joaquin Valley Agricultural Sciences Center » Crop Diseases, Pests and Genetics Research » Research » Publications at this Location » Publication #327299

Research Project: Epidemiology and Management of Pierce's Disease and Other Maladies of Grape

Location: Crop Diseases, Pests and Genetics Research

Title: Quick genome sequencing of “Candidatus Liberibacter” strains by use of Enrichment-Enlargement-Next generation sequencing (EEN)

Author
item Zheng, Z - South China Agricultural University
item Wu, F - South China Agricultural University
item Deng, X - South China Agricultural University
item Chen, Jianchi

Submitted to: American Phytopathological Society Abstracts
Publication Type: Abstract Only
Publication Acceptance Date: 5/19/2016
Publication Date: 7/30/2016
Citation: Zheng, Z., Wu, F., Deng, X., Chen, J. 2016. Quick genome sequencing of “Candidatus Liberibacter” strains by use of Enrichment-Enlargement-Next generation sequencing (EEN). American Phytopathological Society Abstracts. 106:S4.26.

Interpretive Summary:

Technical Abstract: Members of “Candidatus Liberibacter” are associated with several important plant diseases such as citrus Huanglongbing (HLB) and potato zebra chip (ZC) disease. Inability to culture and low titers in infected hosts have been major obstacles for research on these bacteria. The use of whole genome sequence analysis could be an effective means to characterize and study the bacteria. Next Generation Sequencing (NGS) technology with enrichment-enlargement-NGS (EEN) was used for quick genome sequencing of liberibacters. Enrichment includes increasing the ratio of liberibacter DNA to host DNA through using alternative hosts with higher bacterial titer, removal of the highly methylated plant DNA, and selection of samples with low Ct values for liberibacter DNA. Enlargement includes increasing enriched DNA quantity through Multiple Displacement Amplification (MDA) to meet requirements of a sequencing platform. The NGS step includes selection of a sequencing platform and development of specific pipelines/procedures for liberibacters. With EEN, >10 G of sequence data were generated from MiSeq, followed by collection of liberibacter reads using Perl scripts, de novo and/or referenced genome assembling, and sequence annotation. Thus far, seven draft genome sequences have been generated: five HLB liberibacters (China, California, and Florida) and two ZC liberibacters (California) from either plant or psyllid hosts. The EEN procedure has facilitated research on HLB liberibacters recently found in California and could be applied to genome research in other fastidious prokaryotes.