A high-throughout put sequencing SSR markers development and its validation in three developmental stages of fenugreek (Trigonella foenum-graecum L.)

Abstract

Fenugreek (Trigonella foenum-graecum L.) popularly known as “Methi” belongs to family fabaceae and subfamily papilionaceae. Gujarat Methi Variety 2 was selected for transcritome study from the three developmental stages that were vegetative stage (20-30 DAS), reproductive stage (50-60 DAS) and maturity stage (80-90 DAS) which were sequenced using Ion S5 genome sequencing machine. SSR molecular markers were identified individually for each transcript after assembly, by BatchPrimer3 software. A total of 189 primers were developed, out of which 78 from vegetative stage, 64 from reproductive stage and 47 from maturity stage. For validation purpose 30 primers were selected, 10 primers from each developmental stages. The 10 primers for each stage were checked on different stages of cDNA for validation analysis. The primers generated from the vegetative stage were also checked in other two stage of growth. Similarly the primers of reproductive and maturity stages were checked in other two stages of growth. The SSRs containing sequences (Fasta file) which were generated after assembly were also analyzed in NCBI-BLASTn to get the hits for the gene. All the three samples were most related to Medicago trucatula, Arachis duranensis, Arachis hypogea, Arachis ipaensis, Cicer arisetinum, Cajanus cajanand Glycine max.