PN40024, a highly homozygous inbred line originating from ‘Helfensteiner’, was used for T2T genome assembly. In total, 21 Gb (21 024 461 524 bp, ∼42× coverage) HiFi reads were generated by the PacBio platform. For the preliminary assembly, hifiasm was used to assemble the HiFi reads. We then used MUMmer and the 12X.v0 genome version (V. vinifera genome assembly 12X.v0 to order the 38 contigs into 19 chromosomes. The PN_T2T genome size was finally generated (494.87 Mb), being 69 Mb longer than 12X.v0 using the same statistical method. The k-mer metric was used to evaluate genomic homozygosity, estimated at 99.8%. The BUSCO for this genome is up to 98.5%. The PN40024.T2T genome assembly: PN40024.T2T.fa The PN40024.T2T gene annotation: PN40024.gff3 The PN40024.T2T TE annotation: PN40024.TE.gff The PN40024.T2T centromere annotation: PN40024.trf.gff3 The PN40024.T2T protein sequence: PN40024.protein.fa The PN40024.T2T cds sequence: PN40024.cds.fa Comparison of gene annotation among PN_T2T and 12X.v0, 12X.v2, PN40024.v4, PN40024.v4.1: correlation.list Mitochondrial assembly sequence of PN40024: PN_T2T_mit.fa Annotation of mitochondrial assembly for PN40024:PN_T2T_mit.gff3 Chloroplast assembly sequence of PN40024: PN_T2T_chl.fa Annotation of chloroplast assembly for PN40024: PN_T2T_chl.gff3 Citation: Please cite this paper when using the data of PN_T2T for your publications. Xiaoya Shi, Shuo Cao, Xu Wang, Siyang Huang, Yue Wang, Zhongjie Liu, Wenwen Liu, Xiangpeng Leng, Yanling Peng, Nan Wang, Yiwen Wang, Zhiyao Ma, Xiaodong Xu, Fan Zhang, Hui Xue, Haixia Zhong, Yi Wang, Kekun Zhang, Amandine Velt, Komlan Avia, Daniela Holtgräwe, Jérôme Grimplet, José Tomás Matus, Doreen Ware, Xinyu Wu, Haibo Wang, Chonghuai Liu, Yuling Fang, Camille Rustenholz, Zongming Cheng, Hua Xiao, Yongfeng Zhou, The complete reference genome for grapevine (Vitis vinifera L.) genetics and breeding, Horticulture Research, Volume 10, Issue 5, May 2023, uhad061, https://doi.org/10.1093/hr/uhad061