http://informahealthcare.com/mdn ISSN: 1940-1736 (print), 1940-1744 (electronic) Mitochondrial DNA, Early Online: 1–3 ! 2015 Informa UK Ltd. DOI: 10.3109/19401736.2014.1003881

MITOGENOME ANNOUNCEMENT

Mitochondrial genome of the Luchuan pig (Sus scrofa)

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Yanfang Zhang, Zhixun Xie, Xianwen Deng, Zhiqin Xie, Jiabo Liu, Liji Xie, Sisi Luo, Li Huang, Jiaoling Huang, Tingting Zeng, and Sheng Wang Department of Biotechnology, Guangxi Key Laboratory of Animal Vaccines and Diagnostics, Guangxi Veterinary Research Institute, Nanning, Guangxi, PR China Abstract

Keywords

The complete mitochondrial genome sequence of the Luchuan pig was measured by PCR-based methods, primer-walking sequencing, and fragment cloning. The entire mitochondrial genome of the Luchuan pig was identified as a circular molecule consisting of 16,730 bp (GenBank accession number: KP126954). The contents of A, T, C, and G in the mitochondrial genome were found to be 34.65%, 25.80%, 26.19%, and 13.36%, respectively. The complete mitochondrial genome of the Luchuan pig contains a typical structure, including 13 proteincoding genes, two rRNA genes, 22 tRNA genes, and one control region (D-loop region). This complete mitochondrial genome sequence provides essential information in understanding phylogenetic relationships among Sus scrofa domestic mitochondrial genomes.

Genome organization, Luchuan pig, mitochondrial genome

Luchuan pig is derived from the Luchuan county of Yulin city in the south eastern area of Guangxi province in China. The Luchuan pig is one of the kinds of eight famous local varieties in China, and belongs to the national class protected varieties (Huang et al., 2013). The Luchuan pig has some advantages, such as good meat quality, rich nutrition, special growth property, and reproduction performance. Thus, Luchuan pig holds an important position in the market, which is deeply loved by the consumers. At present, with the gradually increasing demand for the number of pork from Luchuan pig, the biological technology and the theory in animal reproduction are needed to apply in the Luchuan pig industry. In addition, the research is developed in reproductive area of Luchuan pig for further improving the breeding performance of Luchuan pig. The excellent resource characteristics should be enhanced, and the protection measurement for Luchuan pig is needed to further do well, for continuously maintaining the predominant status of the Luchuan pig in the national class protected varieties. The most advanced techniques in studying mitochondrial genome provide a unique tool to identify genetically pure bread for the purpose of preservation (Shen et al., 2010). In this report, we describe the organization Luchuan pig’s mitochondrial genomic DNA extracted from its blood using the EasyPure blood Genomic DNA Kit (Beijing TransGen Biotech Co., Ltd., Beijing, China). The complete mitochondrial genome was amplified by 24 pairs of primers designed according to the sequence of the Sus scrofa (GenBank accession number: KF472178). The gel electrophoresis PCR products were purified by using the AxyPrep TM DNA Gel Extraction Kit (Axygen, Hangzhou, China) and submitted to Invitrogen (Guangzhou, Correspondence: Zhixun Xie, Department of Biotechnology, Guangxi Veterinary Research Institute, 51 Youai North Road, Nanning 530001, China. E-mail: [email protected]

History Received 5 December 2014 Accepted 5 December 2014 Published online 29 January 2015

China) for genomic sequencing. The DNA sequence was analyzed using the DNAStar 7.1 software (DNASTAR, Madison, MI) and the base composition and the distribution of the mitochondrial DNA (mtDNA) sequence were analyzed using the tRNA ScanSE1.21 software (http://lowelab.ucsc.edu/tRNA Scan-SE/) (Lowe & Eddy, 1997) and DOGMA software (http://dogma.ccbb.utexas.edu/) (Wyman et al., 2004), respectively. Our research findings have completed the entire mitochondrial genome sequence of Luchuan pig which is 16,730 bp containing 13 protein-coding genes, 22 tRNA genes, two rRNA genes, and one control region (D-loop region), which are similar to those of other avian species in gene arrangement and composition (Xie et al., 2014a). The overall nucleotide composition of A, T, C, and G were found to be 34.65%, 25.80%, 26.19%, and 13.36%, respectively. Most of the genes are encoded on the H-strand (Table 1) and they are similar in structure to the typical mitochondrial genome of vertebrates (Boore, 1999). The Luchuan pig’s entire mitochondrial genome sequence has been deposited in GenBank recently (GenBank accession number: KP126954). The Luchuan pig mitochondrial genes that are not encoded on the H-strand include one protein-coding gene (ND6) and eight tRNA genes (tRNAGln(CAR), tRNAAla, tRNAAsn(AAY), tRNACys, tRNATyr, tRNASer(KCH), tRNAGlu, and tRNAPro(CCN)), which are similar to typical vertebrate mtDNAs (Arnason et al., 2002; Xie et al., 2014b). Of the 13 protein-codon genes for Luchuan pig mitochondria, nine subunits require ATG as the initiation codon, while ND2, ND3, and ND5 gene utilize ATA, ND4L gene use GTG (Table 1). These tRNA genes had four types of stop codons: type 1: AGA for Cyt b; type 2: TAG for ND1 and ND2; type 3: incomplete stop codon ‘T’ for COX2, COX3, ND3, and ND4; type 4: TAA for the others. Type 3 is known to be found in most vertebrates (David et al., 1998; Ojala et al., 1981;

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Table 1. MtDNA genome organization of the Luchuan pig.

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Gene (element) D-loop tRNAPhe 12SrRNA tRNAVal 16SrRNA tRNALeu(YUH) ND1 tRNAIle(AUH) tRNAGln(CAR) tRNAMet ND2 tRNATrp tRNAAla tRNAAsn(AAY) tRNACys tRNATyr COX1 tRNASer(KCH) tRNAAsp COX2 tRNALys ATPase8 ATPase6 COX3 tRNAGly ND3 tRNAArg ND4L ND4 tRNAHis tRNASer(UCR) tRNALeu(YUR) ND5 ND6 tRNAGlu Cyt b tRNAThr tRNAPro(CCN)

Position (from to) Site

Chain Length (bp)

Strand (H/L)

Space (+) Overlap ()

Start

Codons Stop

Anti

1–1294 1295–1364 1365–2325 2326–2393 2394–3963 3964–4038 4041–4997 4996–5064 5062–5134 5136–5205 5206–6249 6248–6315 6322–6389 6391–6465 6498–6563 6563–6628 6630–8174 8187–8248 8254–8321 8322–9009 9010–9076 9078–9281 9239–9919 9919–10,702 10,703–10,771 10,772–11,117 11,119–11,187 11,188–11,484 11,478–12,855 12,856–12,924 12,925–12,983 12,984–13,053 13,054–14,874 14,858–15,385 15,386–15,454 15,459–16,598 16,599–16,666 16,599–16,730

1294 70 961 68 1570 75 957 69 73 70 1044 68 68 75 66 66 1545 71 68 688 67 204 681 784 69 346 69 297 1378 69 59 70 1821 528 69 1140 68 64

H H H H H H H H L H H H L L L L H L H H H H H H H H H H H H H H H L L H H L

0 0 0 0 0 0 2 2 3 +1 0 2 +6 +1 +32 1 +1 +3 +5 0 0 +1 43 1 0 0 +1 0 7 0 0 0 0 17 0 +4 0 0

– – – – – – ATG – – – ATA – – – – – ATG – – ATG – ATG ATG ATG – ATA – GTG ATG – – – ATA ATG – ATG – –

– – – – – – TAG – – – TAG – – – – – TAA – – T– – TAA TAA T– – T– – TAA T– – – – TAA TAA – AGA – –

– GAA – AAC – GAG – GAT TTG CAT – CCA AGC GTT GCA GTA – AGA ATC – CTT – – – GCC – TCT – – ATG TGA TAA – – TTC – GGT TGG

T– represents incomplete stop codons.

Zhang et al., 2014a,b), was probably completed by polyadenlyation of the mRNAs after cleavage of their primary transcripts (Anderson et al., 1981). Positive numbers correspond to the nucleotides separating adjacent genes. Negative numbers indicate overlapping nucleotides. Among all the gene elements (Table 1), 11 overlaps and/or eight spaces were found in the length between 1 and 43 bp. The lengths of the 12S rRNA and 16S rRNA genes are 961 bp and 1570 bp, respectively, they are located between the tRNAPhe and tRNALeu(YUH) genes and separated by the tRNAVal gene. In addition, 22 deduced tRNA genes were found to be distributed in rRNA and protein-coding genes, ranging from 59 to 75 bp in size. The D-loop is a non-coding control region located at 1294 bp position between the tRNAGlu and tRNAPhe genes, where is rich in A and T. However, the length of the control region can potentially fluctuate among individuals due to variation in copy number of the tandem repeat 50 -TACACGTGCG-30 . This region is most likely to show sequence variations. The D-loop contains regulatory elements that control mtDNA replication and transcription.

Declaration of interest The authors report that they have no conflicts of interest. The authors are responsible for the content and writing of the article. This research project

was funded by both the Guangxi Science and Technology Bureau (Grant nos. 1222003-2-4 and 10100014-5) and Guangxi Government Senior Scientist Foundation (Grant no. 2011B020).

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Mitochondrial genome of the Luchuan pig (Sus scrofa).

The complete mitochondrial genome sequence of the Luchuan pig was measured by PCR-based methods, primer-walking sequencing, and fragment cloning. The ...
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