Class I patatin Genes from Indian Potato (Solanum tuberosum L.) Cultivars: Isolation and Partial Characterization

Abstract

The potato (Solanum tuberosum L.) is highly nutritious non-grain starchy food crop belonging to Solanaceae family. It is a short day and C3 plant grown in cool season in temperate regions. Usually the potato cultivars are tetraploid species. The tubers are the sink organs for the storage of starch and other abundant storage proteins. Among the storage proteins of tubers the major soluble glycoprotein is a ~40 kDa protein known as ‘Patatin’ which is encoded by a large multigene family. The sequence variation in the upstream of 5-flanking regions of patatin genes leads to the division of patatin genes in two classes: Class I and Class II. A 22-bp insertion is present only in the 5-UTR of class II patatin genes. Class I patatin genes are expressed at elevated levels mainly in the tubers, whereas class II patatin genes are expressed in roots and tubers but at a very lower level as compared with class I patatin genes. The present study mainly deals on molecular cloning and partial characterization of 5-flanking regions of class I patatin genes of Indian potato cultivars namely Kufri Jyoti (KJ) and Kufri Chipsona-2 (CS-2). A specific set of forward and reverse primer as designed in our laboratory based on the class I patatin gene sequence available in the data base (Accession no. X87216) were used in order to amplify the 5-flanking regions of class I patatin genes. The sizes of the PCR-amplified products were: ~1.5 kb and ~2.0 kb specific to KJ; ~1.0 kb specific to CS-2. The PCR amplified products clearly suggested significant sequence variations in the upstream of 5-flanking regions of patatin genes of different potato cultivars. All these DNA bands were cloned into the SmaI site of a plasmid vector pUC19. For partial characterization, restriction analyses were done to check the presence of the cloned inserts, and PCR was carried out to check the specificity and intactness of the inserts. Based on the sizes of the PCR-amplified products, the following conclusions could be made. The ~1.5 kb DNA band specific to the cv. KJ, in particular, possibly represents a new patatin gene not reported earlier. However, the other DNA bands may show some degree of variations. In order to identify tuber specificity, classification, microheterogeneity and strength of the promoters, sequencing and functional characterization are required.