The Study Of The Insertion/Deletion In Chicken Mitochondrial DNA Genomes

Mitochondria is the essential organelles of eukaryotic animal cells,which might provide 90% energy for vital movement of cell.mitochondrial DNA(mtDNA) is the genetic material that carried by mitochondrial.As the development and perfection of various technology used to research heterogeneity,it has been found that the heteroplasmy of mtDNA is not rare,but a common phenomenon.But the reports that about chicken mtDNA heteroplasmy is little,especially the research on chicken mitochondrial DNA genome insertion/deletion heterogeneity.In this experiment,the leg muscle tissue and blood of Rhode Island Reds,Silkies,Gushi chicken and hybrid chicken were selected,each chicken mtDNA whole genomes were sequenced by Solexa high-throughput sequencing.In the chicken mitochondrial genome,we identified the 49 loci,that the heterogeneity degree were higher than 5 ‰,with 81 types of insertion/deletion.There were 23 loci with 41 types of insertion/deletion in the non-protein coding region and 26 loci with 40 types of insertion/deletion in the protein coding region.We also detected that there were 20 heterogeneity loci which more than 2 insertion/deletion types.The insertion/deletion loci were major focused on D-loop region,16 SrRNA,ND5,COX1 gene,and next were the ND2,ND3,ND6,CYTB gene.But that were not detected in 12 SrRNA、tRNA(22 types),ATP6,ATP8,COX2,COX3,ND1,ND4,ND4 L.The heterogeneity of frequency in non-protein coding region D-loop and 16 srRNA were 0.73% and 0.87% respectively,the heterogeneity of frequency in protein coding region ND5COX1 gene frequency were 0.72% and 0.58% respectively,were higher than that of the mitochondrial genome which was 0.29%.The heterogeneity was abundant with 21 types of insertion/deletion at 9 loci.T-A cloning sequencing experiments have verified the high-throughput sequencing could detect the 0.73% heterogeneity degree.The heterogeneous degree of the heterogeneity loci fluctuated between ~0-36.21%.Most were low heterogeneity loci and the heterogeneity degree were below 10% in 83.87% of those loci.The insertion/deletion in protein coding gene all caused the change of amino acid sequence.We detected the 26 loci with 40 types of insertion/deletion in protein coding gene region.The heterogeneity degree of the protein-coding genes fluctuated between ～0-16.44%.The heterogeneity degree of protein coding genes were low as a whole.The insertion/deletion sequence of the protein coding genes were primarily single-base or two-base.There were 33 single-base and 7 two-base insertion/deletion types,and the 7 two-bases were only detected in the COX1 gene.The ratio of single-base and two-base insertion/deletion were 82.5% and 17.5%respectively.There are five high heterogeneity degree insertion/deletion loci in protein-coding genes,as the 7088 CinsT and 7095 Tins AC in COX1 gene,tha 10948 TinsC in ND3 gene and the 14379 Ains C and 14392 del C in ND5 gene.The amino acid change of COX1 gene were HYS-LH(histidine/tyrosine/serine-leucine/histidine);the Amino acid change of ND5 gene were SNH-ATT(serine/aspartic acid/histidine-alanine/threonine/threonine).There only one base insertion in ND3 gene at the 10948 locus.We predicted that the coding of ND3 gene was interrupted.This phenomenon was common in chicken mitochondria,but mechanism was not unclear.That need our further researchs.The insertion/deletion heterogeneity were non-significant differences in the five groups.That major focused in D-loop region,16 srRNA,COX1 gene and ND5 gene.The heterogeneity loci and heterogeneity degree were non-significant differences in the half-siblings and full-siblings.The significant differences at 2876 Ains C in 098 full-siblings.The half-siblings showed significant differences heterogeneity at 238 Adel G,847 Ains C and 2876 Ains C.The heterogeneity degree at 847 locus in leg muscle and blood of 098-1 and098-2 full-siblings were significant differences,that the heterogeneity degree in leg muscle was higher than that in blood.We identified 36 loci with 63 types of insterion/deletion in the tissues of 068-6 individual.The heterogeneity degree of the all predominant allele of the heterogeneity loci were similar.The floating of heterogeneity degree at 47/703(GinsA)/ 703(GdelA)　loci were slightly larger than other loci.The heterogeneity degree of the abdominal fat tissue was slightly lower than other tissues.Heterogeneity loci and heterogeneity degree at 1 day of age,30 days of age,60 days of age were non-significant difference.This study analyzed the insertion/deletion heterogeneity loci and degree in chicken mtDNA genome in the different groups,full-siblings and half-siblings,different tissues,different ages.That couled supply the theoretical foundation study of mitochondrial.Enriching and improving the mitochondrial genetic theoretics.That also perfected established research foundation for molecular breeding of chicken,the relationship between population genetic structure and system development by mtDNA heteroplasmy.