HiSeq control system (HCS) and real-time analyzer (RTA) are adopted by HiSeq 2000. These two softwares could calculate the number and position of clusters based on their first 20 bases, so the first 20 bases of each se­quencing would decide each sequencing’s output and quality. HiSeq 2000 uses two lasers and four filters to detect four types of nucleotide (A, T, G, and C). The emission spectra of these four kinds of nucleotides have cross-talk, so the images of four nucleotides are not independent and the distribution of bases would affect the quality of sequencing. The standard sequencing output files of the HiSeq 2000 consist of *bcl files, which contain the base calls and quality scores in each cycle. And then it is con­verted into *_qseq. txt files by BCL Converter. The ELAND program of CASAVA (offline software provided by Illumina) is used to match a large number of reads against a genome.

In conclusion, of the three NGS systems described before, the Illu — mina HiSeq 2000 features the biggest output and lowest reagent cost, the SOLiD system has the highest accuracy [11], and the Roche 454 system has the longest read length. Details of three sequencing system are list in Tables 1(a), 1(b), and 1(c).