β-Globin gene mutations reduce or terminate the production of beta globin chains, of which approximately 10% are large deletions within the β-globin gene cluster. Because gene deletion leads to loss of heterozygosity at single nucleotide polymorphism (SNP), a novel method for detecting β-globin gene cluster deletions based on SNP heterozygosity analysis was established in this study. The location range of SNPs was selected according to the breakpoint of β-globin gene cluster deletions. SNPs were screened using bioinformatics analysis and population sequencing data. A novel method which enables genotyping of multiplex SNPs based on tetra-primer ARMS-PCR was designed and optimized. Forty clinical samples were tested in parallel by this method and MLPA to verify the performance of this method for detecting β-globin gene cluster deletion. Six informative SNPs were obtained, achieving heterozygote coverage of 93.3% in normal individuals. Genotyping of six SNPs were successfully integrated into two multiplex tetra-primer ARMS-PCR reactions. The sensitivity, specificity, positive predictive value and negative predictive value of the method for detecting β-globin gene cluster deletion were 100%, 96.30%, 92.86%, and 100%, respectively. This is a simple, cost-effective and novel method for detecting β-globin gene cluster deletions, which may be suitable for use in combination with MLPA for thalassemia molecular testing.