Molecular Marker Mining Of SSR160 Locus In Cotton Mitochondrial Genome And Its Effect On The Stability Of AtpA MRNA

Cotton is an important economic crop in our country.Gossypium can be divided into 8 diploid chromosomes A,B,C,D,E,F,G and K,and AD heterotetraploid chromosomes.DNA molecular marker technology has promoted the development of cotton breeding in my country,but there are few studies on DNA molecular markers in cotton mitochondrial genome.The previous study found that there was an SSR site which was located 160 bp downstream of the stop codon of the atp A gene in the cotton mitochondrial genome.We named it SSR160.This site is significantly polymorphic between the male sterile line and the maintainer line of upland cotton.In this study,the sequence of SSR160 in 29 species of diploid and tetraploid cotton were cloned and sequenced,and their sequence polymorphisms were analyzed.And the relationship between the length of the site sequence and the stability of mRNA was analyzed.The results were as follows:1)Using MISA software,a total of 107-140 SSR loci were detected in the mitochondrial genomes of 7 cotton species including G.arboreum,G.thurberi,G.davidsonii,G.raimondii,G.trilobum,G.hirsutum and G.barbadense.The total length of SSR sites is 1 271 bp-1 656 bp,and their average length is 11.80 bp-11.97 bp;the density of SSR occurrence is 1.90-2.44 bp/kb.2)There is a compound short repeat sequence at the SSR160 locus:（TAA）m（TA）n.Among these 29 cotton species,A,B and AD are all（TAA）₇（TA）₆;D cotton has two forms of（TAA）₄（TA）₃ and（TAA）₃（TA）₂;C type cotton is（TAA）₅（TA）₅and（TAA）₅（TA）₆;E type cotton is（TAA）₄（TA）₃and（TAA）₄（TA）₄;G cotton has two forms:（TAA）₅（TA）₆ and（TAA）₄（TA）₄.3)Twenty-six SNP sites have been discovered.Using these short repeat sequences and SNP sites,these 29 cotton species can be divided into 17 different groups:（1）:G.arboreum（A₂）,G.africanum(A_1-2),G.anomalum（B₁）;（2）:G.sturtianum（C₁）;（3）:G.robinsonii（C₂）;（4）:G.thurberi（D₁）,G.trilobum（D₈）,G.stocksii（E₁）;（5）:G.harknessii（D₂）,G.turneri(D₁₀);（6）:G.klotzschianum(D_3-K);（7）:G.davidsonii(D_3-d);（8）:G.aridum（D₄）,G.gossypioides（D₆）,G.laxum（D₉）,G.lobatum（D₇）,G.schwendimanii(D₁₁);（9）:G.armourianum(D_2-1);（10）:G.somalense（E₂）,G.nelsonii（G）,G.australe（G）;（11）:G.bickii（G₁）;（12）:G.hirsutum（AD₁）;（13）:G.barbadense（AD₂）;（14）:G.tomentosum（AD₃）,G.darwinii（AD₅）;（15）:G.mustillinum（AD₄）;（16）:G.nandewarense（C₁）;（17）:G.raimondii（D₅）.4)Northern blot was used to detect the difference in the abundance of the full-length atp A gene mRNA transcripts of the upland cotton cytoplasmic male sterile line P30A and the maintainer line P30B.The results showed that atp A gene transcript abundance had no significant difference between the sterile line and the maintainer line.This indicates that the deletion of the sequence in the SSR160 region may not reduce the stability of atp A mRNA in the sterile line.The SSR160 marker is of great significance for understanding the genetic diversity of cotton germplasm resources and expanding the basis of cotton germplasm;comparing the genetic relationship between 29 types of cotton also provides a biological basis for cotton hybrid breeding.At the same time,we did not find significant differences on the abundance between P30A and P30B,indicating that the polymorphism of SSR160 may not have a significant impact on the stability of atp A gene mRNA.The result deepens the understanding of gene expression in mitochondrial genome of cotton.