# Identification of Seasonal Honey Based on Quantitative Detection of Typical Pollen DNA
## 1 Introduction  
Honey has been widely consumed as a safe product by humans owing to its appreciable taste and health benefits; however, it has been targeted for economic fraud [1]. In general, monofloral honey is produced from the nectar of single predominant botanical species [2; 3; 4]. Some monofloral honey are expensive because of their unique and medicinal properties, and it is conveniently targeted for adulteration by incorrect labeling or mixing with other cheaper honey samples [5; 6; 7; 8]. Honey adulteration involves tricking consumers, leading to unfair competition in the market share [9].  
Pollen that remains in honey has been used as an indicator of the floral origin of honey as well as the harvest season. The melissopalynology method was used to identify botanical species based on pollen morphology observed using light microscopy [10; 11; 12]. However, this low-cost method must be conducted by a botanical specialist and is time-consuming to observe and distinguish all the pollens by microscopy [13]. In addition, the reference data for species identification using pollen is scarce [14]. Therefore, an alternative method based on molecular detection was developed to authenticate the botanical origin of honey [15; 16; 17]. The method was performed using specific primer for PCR amplification and the reference sequences from databank for species identification. Consequently, detection of specific plant species based on the DNA that is isolated from the pollen in honey is considered as a reliable method for the accurate determination of honey's origin. In addition, several samples can be analyzed in one PCR performance by a molecular technician. Therefore, time spent for species identification in collected pollens can be saved. Several methodshave been developed to improve DNA isolation from pollen in honey, such as breaking the cell wall of collected pollens by ultrasonication or vortexing with glass beads and followed by DNA isolation using Nucleospin, DNeasy, Wizard, or CTAB method [18; 19].  
In South Korea, beekeeping has important contribution to the national economy. The number of honey bee colony was 1,756,993, and annual value of honey product was estimated with around USD 535 million in 2013 [20; 21]. The number of colonies increased to 2,697,842 in 2020 and the value of beekeeping industry was estimated with approximately USD 5.5 billion [21; 22]. Around 555 honey plant species were identified in the country, of which cherry blossom (_Prunus_ sp.), false acacia (_Robinia pseudoacacia_), chestnut (_Castanea_ sp.), and aralia (_Kalopanax_ sp.) flower were identified to bloom within two weeks and majorly provide nectar and pollen to honey bees to produce corresponding monoffol honey in April to July, respectively [23; 24; 25]. Therefore, the monoffolared honey of the four plant species are popular in the country, in which false acacia honey accounts for 70% of honey production in the country annually [23; 24].  
In the present study, we attempted to develop a method to authenticate the origin of monoffol honey and the season during which it was produced. The quantity of DNA in the natural honey samples was determined and compared to identify the major source of nectar in each sample.