Identify the genus origin of animal glue used in Chinese historical mortars using a new DNA mini-barcoding method

In historic buildings and paintings, animal glue was one of the first and most used organic binders. The identification of these binders would provide a clue for the life of the ancient human beings. However, the precise detection of the clue is challenging as the amount of the glue is very low and always mixed with other impurities. Previously, the ELISA method was successfully used in the identification of the glue in family level. However, the method can not tell the species of the glue. For example, it can not known whether the glue was from cattle or sheep using ELISA methods. This study provides a novel biological technique to identify the species of animal glue used as ancient binders. Hence, the accurate identification results at the species level can be obtained by DNA barcoding method with low detection limits. Moreover, ten animal glues belong to different species can be identified using a pair of universal primer at one time. These species include pigs, cattle, buffalo, goats, sheep, camels, deer, dogs, horses, and rabbits. Actually, the species of animal glue used in mortars obtained from three cultural heritages, the Neolithic Yulin City in Shaanxi Province, the Forbidden City in Beijing, and the Confucian Temple in Qufu, Shandong Province, China, were successfully identified. It was demonstrated that certain animals were not only consumed as food resource at that time, but their leftover bones, skins, and other parts were also utilized to produce animal glue. Additionally, our findings is consistent with the records in ancient books, which documented the species of animals frequently used in the manufacturing of animal glue. This is the first report using DNA barcoding method for the identification of animal glue in historic mortar. The identification of animal glue's species can not only help archaeologists understanding the agriculture and animal husbandry in certain area, but also can help people find the suitable materials for restoration and conservation of cultural heritage.