Shrimp aquaculture is facing a serious disease, acute hepatopancreatic necrosis disease (AHPND), caused by Vibrio paraheamolyticus (Vp_APHND). For sustainable shrimp aquaculture, massive losses of shrimp infected with Vp_APHND must be prevented. Research and selection of shrimp tolerant to Vp_APHND infection is a sustainable approach to reducing the risk of AHPND. This study focused on the identification and development of potential DNA markers associated with AHPND using DArT sequencing (DArTSeq) and a genome-wide association study. Three populations of post-larval Litopenaeus vannamei were immersed in Vp_APHND to collect susceptible (D) and tolerant (S) samples. The 45 D and 48 S shrimp had their genotypes analyzed using DArTSeq. A total of 108,983 SNPs and 17,212 InDels were obtained from the DArTseq data, while the biallelic 516 SNPs and 2293 InDels were finally filtered with PIC < 0.1, MAF < 0.05, and a call rate ≥ 80%. The filtered variants were analyzed for their association with AHPND tolerance. Although there were no significantly associated SNPs and InDels above the Bonferroni correction threshold, candidate variants, four SNPs and 17 InDels corresponding to p < 0.01, were provided for further validation of the AHPND tolerance trait. The candidate SNPs are located on an exon of the zinc finger protein 239-like gene, an intron of an uncharacterized gene, and in intergenic regions. Most of the candidate InDels are in the intergenic regions, with fewer in the intronic and exonic regions. This study provides information on SNPs and InDels for white shrimp. These markers will support the variant database of shrimp and be useful in shrimp aquaculture for breeding selection.

Taq polymerase system is one of the key success factors for genotyping by using semi-random marker types. Start codon targeted (SCoT) and sequence-related amplified polymorphism (SRAP) are semi-random markers that amplify more than one target at a time, like in multiplex PCR. Here, we compared the performance of five multiplex PCR kits (QIAGEN^®, Thermo Scientific™, New England Biolabs, KAPA Biosystem and iNtRON Biotechnology) and Taq DNA polymerase from Vivantis as a representative common Taq polymerase. The PCR using three SCoT and three SRAP markers was performed in six plants, which were different in genome size, including three monocotyledons (rice, oil palm and onion) and three dicotyledons (holy basil, chilli pepper and eggplant). The overall results demonstrated consistent high performance of the Taq polymerase system with the multiplex PCR kit from KAPA Biosystem. Considered by plant, the multiplex PCR kit from KAPA Biosystem showed the best performance in holy basil, eggplant, oil palm and chilli pepper, whereas in rice, the Taq DNA polymerase from Vivantis generated better results than the KAPA Biosystem. In onion, multiplex PCR kit from Thermo Scientific™ showed the best performance among those tested, even if it did not perform well in the other plans. Different Taq polymerase systems might generate different band patterns, therefore using the same Taq polymerase system within one experimental dataset is recommended. The multiplex PCR kit from KAPA Biosystem is the best starting choice.

Insertion-deletion (InDel) markers are co-dominant, relatively abundant and practical for agarose gel genotyping. InDel polymorphism usually affects gene functions. Nucleotide sequences of durian (Durio zibethinus) are available, but InDel makers have not been well established. This study aimed to develop drought-related gene-based InDel markers for durian through bioinformatic analysis of RNA-Seq datasets. The polymorphism of the markers was verified in 24 durian genotypes local to Thailand. Bioinformatic analysis indicated 497 InDel loci having lengths more than 9 bp. To evaluate these InDel markers, 15 InDel loci were selected. Nine markers were successfully amplified a clear polymorphic band pattern on 2% agarose gel. The polymorphic information content (PIC) of these nine markers ranged from 0.1103 to 0.5808. The genetic distance between the 24 genotypes ranged from 0.222 to 0.889. The phylogeny based on the nine InDel loci distinguished the 24 genotypes, and divided samples into four groups. This set of gene-based InDel markers on putative drought responsive genes will be useful for genetic studies.