ABBS 2004,36(6)::High Throughput SNP Genotyping with Two Mini-sequencing Assays

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ISSN 0582-9879 Acta Biochim et Biophysica Sinica 2004, 36(6):379-384 CN 31-1300/Q

High Throughput SNP Genotyping with Two Mini-sequencing Assays

Chunqing LUO^1,2#, Libin DENG^1#, and Changqing ZENG^1,2*

¹The Institute of Genetic and Developmental Biology, the Chinese Academy of Sciences, Beijing 100101, China;

²Beijing Genomics Institute, the Chinese Academy of Sciences, Beijing 101300, China

Abstract Two mini-sequencing methods, FP-TDI (template-directed dye-terminator incorporation with fluorescence-polarization) and MassArray (matrix assisted laser desorption ionization time of flight detection mass spectrometry), were optimized. A numeric standard was introduced to evaluate the SNP scoring quality of FP-TDI assay, thus made the optimization work easier. At the same time, using multi-PCR technology, 8-plex genotyping of MassArray assay was successfully carried out, some softwares were developed and the data process of MassArray was highly automated. Then these two methods were applied to high throughput SNP genotyping, the accuracy, efficiency and robustness were compared. The result shows FP-TDI is more sensitive to the concentration of SNPprimer and PCR product, as well as extension cycles, the SNPprimer length of FP-TDI should be 24-30 bp long, whereas MassArray assay prefers to be as short as only 16 bp. Altogether 6440 SNP sites of human chromosome 3 were genotyped in a sample of 90 individuals, 4792 sites by FP-TDI assay and 1648 sites by MassArray assay, the success rates of FP-TDI and MassArray were 67.7% and 93.6% respectively. The throughput of MassArray was higher than FP-TDI, and the cost of MassArray was lower, MassArray was more suitable for high throughput SNP genotyping.

Key words SNP genotyping; FP-TDI; MassArray; high throughput

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Received: March 8, 2004 Accepted: April 15, 2004

This work was supported by a grant from the Major State Basic Research Development Program of China (No. G1999055901)

*Corresponding author: Tel, 86-10-65296457; Fax, 86-10-65296466; Email, [email protected]