Recovery of DNA signatures from historical herbarium specimens using chloroplast and nuclear barcodes

Abstract

Expert-verified curated collections of herbarium specimens are invaluable assets of plant biodiversity. Several historical specimens of rare and endangered taxa in herbaria face difficulty in identifying the morphology. The emerging DNA-based technology recovers genetic data from preserved herbarium are efficiently used for plant identification complementing taxonomy. Therefore, the current study utilized DNA barcodes from chloroplast (rbcL, trnH-psbA) and nuclear (ITS2) genomes to identify chronologically preserved specimens of endemic and threatened taxa. Fifteen herbarium accessions belonged to the 19th, 20th, and 21st centuries representing eight taxa studied as test samples by comparing with three control samples. Extraction of DNA from 16 to 140-year-old herbarium showed positive results using our standardised isolation protocol. The PCR amplification was successful using rbcL DNA barcode in all the samples but the trnH-psbA and ITS2 markers amplified only two species. The DNA sequence recovery from ∼100 old specimens yielded short fragments ranging between 100 bp and 150 bp. The DNA sequence was successfully recovered from the sixty and twenty-year-old herbarium specimens of Ischaemum santapaui and Lepidagathis lutea, respectively. The presence of mononucleotide repeats affected the sequence recoverability in trnH-psbA and ITS2 markers. Our results indicated that the increased rate of DNA degradation, fragmentation, and mixing of microbial DNA in the specimen during long-term storage significantly affected the PCR amplification in preserved herbarium specimens. This study highlighted the application of DNA barcoding in unmasking the trove of genetic diversity in the present and historical herbarium collections.