| In this experimentation, random amplified polymorphic DNA (RAPD) techniquewas used to analyze the genetic relationship of seven cultivars, twelve wild speciesand six relative of Hibiscus cannabinus L.. The extraction methodology of genomicDNA of Hibiscus cannabinus L. was explored and the influential parameters wereoptimized. On the basis of them, RAPD was used to investigate the geneticpolymorphisms among the twenty-four breeds. And both the genetic relationships andthe genetic differences of them were to be researched, furthermore, the Kenaf'scradleland and its transmit route were also discussed. Results were as follows: 1. To save time and reduce workload, a steady, efficient, and applicableextraction method of Kenaf's seeds genomic DNA for RAPD analysis, which isimprove greatly on the general method, was explored. and the key points of themethod are triturating after adding extractive buffer(SDS) and shaking intensivelyafter adding CIAA. The genomic DNA samples, prepared from seeds of Hibiscuscannabinus L. with aforementioned modified method were to be used as templates forpolymerase chain reaction (PCR) and very suitable for RAPD analysis. 2. Steady reaction system is the key point to RAPD analysis, in order to reachthis, The influential factors such as templates DNA, dNTPs, MgCl2, primers, Taqpolymerase and so on were studied and experimental parameters were optimized.They were as follows: 25μl reaction system containing 1×buffer, 2.0mmol/LMgCl2,200μmol/L dNTPs, 0.6μmol/L primers, 45ng template, 1.0U Taq polymerase. Theoptimal amplification program was as follows: 5min at 94℃, followed by 94℃ for30sec, 38℃ for 45sec, 72℃ for 1min30sec, 40 cycles and final extension time of 72℃ for 7min.This optimized experiment conditions could obtain reproducible andcredible results. 3. Twenty-one primers selected from one seventy primers amplified 146 RAPDfragments and 118 bands of them showed polymorphism, which occupied 85%. Thelength of most amplified fragments ranged from 0.2 to 1.5kb. The dendrogram wasconstructed by using DPS software and Unweight Pair Group Method with ArithmeticMean based on their genetic diversity calculated from RAPD data. It was concludedthat the twenty-four breeds could be classified into three groups-Kenaf's cultivars,Kenaf's wild breeds and Kenaf's relatives, it relatively accorded with knowngenetic relationships.. 4. Using RAPD technique analysis , the genetic relationships among thetwenty-four breeds was investigated, and the results were as follows: There is greatgenetic difference among those specimen and that in spite of H004,H018 are wildstirp, they still be classified into the group of Kenaf's cultivars, shows that Thegenetic difference between H004,H018 and Kenaf's cultivars were relatively closercomparing with the other wild breeds. In addition to the great genetic differenceamong those specimen, the genetic diversity among Kenaf's cultivars isn't as rich asthe morphological analysis shows, while the genetic diversity among Kenaf's wildgermplasm is richer than its morphological analysis shows. Accordingly, the researchof Hibiscus cannabinus L. on molecular level is extraordinary important. 5. Base on the study of genetic relationships of the twenty-four breeds, theKenaf's cradleland and its transmit or evolution route were explored in thisexperiment. The results of RAPD analysis show that Kenaf's wild breeds wasclassified between cultivars and its relatives, besides, two wild breeds H004,H018was also classified into the group of its cultivars. It indicated that genetic relationshipsbetween Kenaf's wild breeds and its cultivars closer than that between wild breedsand Kenaf's relatives, meanwhile, cultivars and relatives keep a certain geneticrelationships through wild breeds. The results support the hypothesis that Hibiscuscannabinus L. rise from Africa in some extent. According to results obtained byRAPD analysis mentioned above, it is safe to say that RAPD technique is suitable tostudy the Kenaf's cradleland and its transmit or evolution route.
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