| Banana(Musa spp.) is one of the most important tropical and subtropical fruits, and is the fourth largest food crop after rice, wheat and maize. However, banana cultivation is facing biotic and abiotic stress, so fostering anti-pest, wide adaptability, high yield banana varieties, is the only way to fundamentally solve such problems.The most of cultivars are originated from intra- and interspecific hybridizations between seed bearing subspecies of M. acuminata and M. balbisiana. So Musa germplasm resources mostly contain A genome and B genome, and they have various levels of ploidy and genomic constitutions. Genome composition has played an important role in classification of bananas and genetic improvement. It is difficult to identify the genome type by the external morphological characteristics. And it is easily affected by cultivated conditions and environmental factors.In this study, we used molecular marker of ITS-PCR-RFLP and copia –IRAP to determine the genomic constitution and used flow cytometry to determine ploidy quickly.The study identified 232 parts banana resources. By this experiment to identify and correct some mistakes of genome type from the collection abroad that they were dentify by morphological characteristics; to explore a number of AA, AB and BB types of precious resources to provide the genetic basis of cross breeding parents choice.,and to provide the basis for the identification of molecular breeding work. Also we found through experiments using IRAP further study of genetic diversity of banana germplasm.The main results are as follows:1、The method for the ploidy identification of Musa with flow cytometry was explored and established. Zea mays is the inter standard(2C=5.43pg). Using this method, 183 accessions were detected, involving cultivars, wild and hybrids. The result shou that183 accessions include the 69 diploid accessions, 91 triploids, 23 tetraploid accessions and 97.1% of the results are consistent with the reported results(Dwarf French Plantain、IRFA 908、NBB 11 are inconsistent with the reported results). 79 accessions without reported result are identical with morphological characteristics. The 2C values were estimated, the diploid accessions range from 0.92 pg to 1.28 pg, the triploids vary from 1.50 pg to 1.80 pg, the tetraploid accessions range from 1.97 pg to 2.37 pg. The CVs of the Zea mays are ranging from 1.5% to 4%, whereas the CVs of Musa are varying from 2% to 8%.2、The method for the ploidy identification of sexual hybridization offspring with chromosome counting was explored and established. For the identification of sexual hybridization offspring, flow cytometry can not be clearly identified, so chromosome counting was used to detect its ploidy directly. In this study, a protoplast dropping technique and DAPI as the stain was been used to find the well-spread metaphases for chromosome counting easily and to avoid any problems of small chromosomes and polyploidy on squash preparations. In the result of 6 habrids, Z-33 is triploid, while others are tetraploid.3、The genome constructions of 232 accessions were identified by molecular marker, ITS-PCR- RFLP and copia-IRAP. The ITS sequence, about 700 bp, was amplified by specific primers. The specific bands that characterized of A genome and/or B genome will be present after digestion for ITS fragments with Rsa I. The fragments of 530 bp specific for A genome, 350 and 180 bp specific for B genome. The accessions containing the B genome would be amplificated by copia-IRAP with 420 bp fragment to determin the number of B genome. Two bands about 200 bp reprent the accession containing two or more B genomes; the fragments of 300 bp and 100 bp reprent one B genome. The results show that 96% of the results are consistent with the reported results in 229 accessions. Eight accessions are inconsistent with the reported results, including YJ086(AAAA)、YJ087(AAA)、YJ181(only A genome)、YJ203(no A or B)、YJ193(A and two or more B)、YJ208(A and two or more B)、YJ211(only A genome)、YJ048(A and B genomes). |
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