Cloning And Expression Analysis Of Atp1Gene In Cytoplasmic-nuclear Male Sterile Line And It’s Maintainer Line Of Hibiscus Cannabinus L.

Kenaf(Hibiscus cannabinus L.) is an annual fast-growing fiber crop of malvaceae hibiscus with betteradaptability.In addition to the traditional application,nowadays we are mainly looking for itsmultifunctional development,such as making paper,wall cloth,linen mulch,forage,and so on.So kenaf isconsidered to be one of the most potential crop of21st century for of multipurpose.The utilization of heterosis is one of the effective ways to improve yield of kenaf.And the discoveryof male sterile plants of kenaf have provided a new way for the utilization of heterosis,which could alsobe effectively avoided the disadvantages of chemical and artificial emasculation in heterosisutilization.With technological development of modern molecular biology,it will be of great significancefor hybrid seed production to get a further understanding the mechanism of male sterility in kenaf.Use the cytoplasmic-nuclear male sterile line and its maintainer line of kenaf which got from the malesterile plant discoverd in Hainan through backcross breeding as the research object,the method ofhomology cloning and RACE,the atp1gene in maintainer line was first cloned and sequence wasanalyzed.Sequence homology analysis was carried on among different species.Then atp1genes werecloned from DNA and cDNA of male sterile line and maintainer line,and the differences between malesterile line and maintainer line was detected after the cloning.Finally,compared with the expressiondifferences of atp1gene among different issues of sterile line and maintainer line,we will move forwardto confirm the relationship between atp1gene and the male sterility.The main results of this study are asfollows:(1) This is the first time that the encoding gene of ATP synthase alpha subunit-atp1was cloned inkenaf.The full length of cDNA is2315bp.The biggest open reading frame predicted is1524bp.There isthe initiation codon ATG at the216th bp of5’ end and terminaton codon TGA at the1737th bp of3’end.And also poly(A) tail is existed at3’ end.The CDS region encodes507amino acid residues withpredicted molecular weight of55.1x103D and a isoelectric point of6.23.Gene sequence has beensubmitted to GenBank with a sequence number of KC713790.Through the homology analysis,we canget that the atp1gene in kenaf shares high homologous with some other species which reach up to94%-98%.And the phylogenetic analysis showed that atp1gene in kenaf had the closest reltionship withthat in soybean.(2) atp1gene was amplified in kenaf male sterile line and its maintainer line respectively on the levelof DNA and RNA.After sequence aligning,we found that the sequences of atp1gene were all samebetween kenaf male sterile line and its maintainer line.Semi-RT-PCR analysis result indicated atp1geneexpressed in root,stem,leaf,petal,anther and pistil.And the expression of atp1in male sterile line waslower in leaf,anther than the maintainer line.