An evaluation of multiple annealing and looping based genome amplification using a synthetic bacterial community
- PDF / 672,221 Bytes
- 6 Pages / 595 x 842 pts (A4) Page_size
- 117 Downloads / 168 Views
An evaluation of multiple annealing and looping based genome amplification using a synthetic bacterial community WANG Yong1, 2†, GAO Zhaoming1, 2†, XU Ying2, 3, LI Guangyu4, HE Lisheng1, 2, QIAN Peiyuan2* 1 Institute of Deep Sea Science and Engineering, Chinese Academy of Sciences (CAS), Sanya 572000, China 2 Division of Life Science, Hong Kong University of Science and Technology, Hong Kong, China 3 School of Life Science, Shenzhen University, Shenzhen 518000, China 4 Key Laboratory of Marine Biogenetic Resources, Third Institute of Oceanography, State Oceanic Administration,
Xiamen 361005, China Received 20 August 2014; accepted 26 December 2014 ©The Chinese Society of Oceanography and Springer-Verlag Berlin Heidelberg 2016
Abstract
The low biomass in environmental samples is a major challenge for microbial metagenomic studies. The amplification of a genomic DNA was frequently applied to meeting the minimum requirement of the DNA for a high-throughput next-generation-sequencing technology. Using a synthetic bacterial community, the amplification efficiency of the Multiple Annealing and Looping Based Amplification Cycles (MALBAC) kit that is originally developed to amplify the single-cell genomic DNA of mammalian organisms is examined. The DNA template of 10 pg in each reaction of the MALBAC amplification may generate enough DNA for Illumina sequencing. Using 10 pg and 100 pg templates for each reaction set, the MALBAC kit shows a stable and homogeneous amplification as indicated by the highly consistent coverage of the reads from the two amplified samples on the contigs assembled by the original unamplified sample. Although GenomePlex whole genome amplification kit allows one to generate enough DNA using 100 pg of template in each reaction, the minority of the mixed bacterial species is not linearly amplified. For both of the kits, the GC-rich regions of the genomic DNA are not efficiently amplified as suggested by the low coverage of the contigs with the high GC content. The high efficiency of the MALBAC kit is supported for the amplification of environmental microbial DNA samples, and the concerns on its application are also raised to bacterial species with the high GC content. Key words: bacterial DNA, MALBAC, metagenome amplification Citation: Wang Yong, Gao Zhaoming, Xu Ying, Li Guangyu, He Lisheng, Qian Peiyuan. 2016. An evaluation of multiple annealing and looping based genome amplification using a synthetic bacterial community. Acta Oceanologica Sinica, 35(2): 131–136, doi: 10.1007/s13131-015-0781-x
1 Introduction High throughput next-generation-sequencing technologies have greatly facilitated research on environmental microbes, as they enable the direct assessment of microbial genomes and bypass the isolation and cultivation procedures in the laboratory (Logares et al., 2012). Given the advantage in sequencing efficiency, a fundamental limitation is the low biomass of samples, such as those obtained from minerals and hydrothermal vents. Amplification technologies were developed to cope with this probl
Data Loading...
