GENETIC VARIATION ANALYSIS ON PEPPER YELLOW LEAF CURL VIRUS (PYLCV) FROM SEVERAL CHILI (Capsicum annuum) FARM CENTERS AND THE USE FOR VIRUS MOVEMENT PREDICTION IN WEST JAVA CHILI FARM

• Main Author: Ayu Kurniasari, Diah
• Format: Theses
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/43485
• Institution: Institut Teknologi Bandung

Pepper Yellow Leaf Curl Virus (PYLCV) is a Begomovirus that attacks chili plants which mediated by Bemisia tabaci. The damage caused by an adult viruliferous Bemisia tabaci in chili farm can reach 30-50%. The effectiveness of PYLCV transmission is influenced by the AV1 gene which is a coat protein encoding gene that plays a role in the encoding of viral ssDNA, transmission into plants, and vector specificity. Identification of AV1 PYLCV gene variation from a Bemisia tabaci can provide information about the level of difficulty in chilli cultivation land management. The higher the variation of AV1 PYLCV gene in the chilli cultivation field, the more difficult to control. The purpose of this study was to identify variations of AV1 genes taken from the body of Bemisia tabaci in chili cultivation centers, by making universal primers AV1 PYLCV (Primer AYC), and phylogenetic analysis of AV1 genes. The method used includes designing of a universal primer based on 90 AV1 genes taken from several countries in Asia (NCBI), Bemisia tabaci sampling in 9 districts of chilli cultivation centers in West Java, isolation of viral DNA taken from a Bemisia tabaci which randomly picked from each sub-district, AV1 gene amplification using PCR with AYC primers (designed), purification and ligation of AV1 gene into pGEM-T Easy, transformation of ligation results to E.coli DH5?, blue-white selection and white colony picking for colony PCR with M13, plasmid isolation with alkaline lysis, confirmation the results of plasmid isolation by PCR using M13 primers, sequencing using M13 primers, data sequencing processing, and phylogenetic analysis on nucleotides and amino acids AV1 gene of PYLCV using Neighbor Joining algorithm and 1000x bootstrap. The results of study obtained are primer AYC forward: 5’-GGMCCDTGTAAGGTCCARTCYTT-3', and primer AYC reverse: 5'- GCRTGVGTACAYGCCATRTACA-3'. In addition, phylogenetic analysis showed the presence of one large group of AV1-PYLCV genes in West Java which consist with 2 subgroups, and one outgroup. Subgroup I (chili monoculture) consists of Cigalontang, Kadipaten, Arjasari, and Pasir Wangi, while subgroup II (chili-tomato polyculture) consists of Ciwidey, Cikajang, Cigedug, and Sukaraja. Campaka Mulya has the most different variation of AV1 gene, hence it is categorized as an outgroup in the phylogenetic tree of AV1 gene, but the most variation AV1 gene come from sampel Pasir Wangi. In conclusion obtained is that the AYC primers can be used to determine AV1 gene variations, which could be influenced by cropping systems and farm conditions. The general conclusion from this study is AV1 PYLCV gene variation in monoculture chilli farming and polyculture chilli farming is quite different, but it is still in the same group. This information will be useful for determining PYLCV control techniques in West Java.