| Cytoplasmic male sterility (CMS) is determined by nuclear-cytoplasmic interactions. Because CMS has an important economic relevance for the generation of hybrid and offers an attractive opportunity for exploring the mechanism how the nuclear genes regulate mitochondrial genes in multicellular organisms, it has received considerable attention during the past years both from breeders and biologists.Now lots of studies indicated that cytoplasmic DNA was associated with CMS, and many CMS-related DNA fragments from mitochondrial (mt) DNA and chloroplast (cp) DNA have been obtained and sequenced. But most of them from different plant species did not show any degree of sequence similarity to each other. Even in cases that more than one type of CMS was found in one species, the DNA fragments possibly involved in conferring the trait appear to be unrelated. This means the hypothesis of the mechanism of CMS resulting from one case does not fit for another. Obviously, identifying common gene(s)-associated with CMS from different plant species or CMS types is very necessary.With this idea in mind, ten CMS lines of sorghum (Sorghum bicolor), seven fertile lines, and two fertility-restored Fl hybrids were chosen in this study. The ten CMS lines have AI, Aa, A3, A4, AS, Ae, and 9E cytoplasms, respectively, which include the main sterilizing cytoplasm types used in breeding of sorghum. The seven fertile lines include 4 maintainers and 3 restorer lines with different nuclear backgrounds. Among the CMS lines and maintainers, Lines AITx398~A6Tx398 and 9ETx398 are alloplasm and isonuclear to line Tx398, lines AiTx623 and A2V4 are isoplasm and allonuclear to lines A]Tx398 and A2Tx398, respectively, and lines AiTx623 and A2V4 are allonuclear and allonuclear to A3 Jinliang 5. Moreover, there were complicated maintaining-restoring relationships among AI, A2, and A3 systems. Comparison study of these CMS plants from different ways would lead to identification of the DNA fragments specific to the cytoplasm or the nuclear DNA of the CMS lines.At first, total DNA of two CMS lines AiTx623 and A2V4 were compared employing RAPD (Random Amplified Polymorphic DNA) with correspondingmaintainers, Tx623 and ?. Five random sequence 10 mer primers were selected from total of near 200 primers because of their consistent amplification of five DNA fragments, designed SAP-069so, SAAU-0270o, SAY-17220o, SAY-19230o, and SAR-Ohsoo, specific to the male sterile (S) cytoplasm or the male fertile (N) cytoplasm. Then, to investigate whether or not the five fragments were still associated with the N- or S- cytoplasm among more CMS materials, the total DNA from other five fertile lines, four CMS lines, and two fertility-restored lines were further subjected to RAPD analysis with the selected five primers.Two of the above five fragments, SAY-1722oo and SAR-0125oo appeared both in the amplification of plants with N cytoplasm or S cytoplasm, so the two segments were not specific to N or S cytoplasm. In contrast, fragments SAP-069Jo presented only in the PCR patterns of sorghums with S cytoplasm, and SAAU-0270o and SAY-1923oo appeared only in the amplification of plants with N cytoplasm, no exception was observed. Those results revealed that the association between SAP-06950 and male-sterile cytoplasm, and between SAAU-027oo, SAY-1923oo and male-fertile cytoplasm, implying that the three fragments might be related to CMS of sorghum.So, the three fragments were retrieved by an approach discovered by us from the gel, then cloned into vector pT7T3 18U/19U and sequenced.1. Fragment SAP-069so This fragment was a new gene of mtDNA, which showed homology to nuclear DNA and revealed differences between the nuclear DNA of CMS line and maintainer.The total length of this fragment was 967bp Analysis of the GenBank nucleotides sequence database with "blastn" found no registered sequence showing significant homology to it, indicating it's a new sequence. An ORF beginning with ATG was identified by GENESCAN. This ORF, which encodes 223 ami...
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