ISSR Analysis Of The Germplasm Resources Genetic Diversity Of Luffa

Luffa belonged to Cucurbitaceae, Luffa Mill.It was generally divided into two cultivars ,Luffa cylindrical (L.) Roem and Luffa (Luffa acutangula (L.) Roxb. Luffa was widely cultivated in china and famous for it's tender fruit and delicious flavor, was one of the main vegetables in summer. Existing reserachs were mainly focused on the cultivation, conventional breeding, extraction and analysis of medicinal ingredients and processing technology ,but the study of luffa molecular biology was relatively few. The test aimed at a preliminary discussion of the genetic diversity of luffa and the early work of molecular level breeding . 30 domestic varieties luffa materials were collected for testing. ISSR-PCR molecular marker technology was applied to explore the classification of luffa and genetic diversity. The main conclusions were as follows:⑴Because of luffa leaves rich in phenolic compounds and polysaccharides which lead to DNA extraction and purification obstacles, the traditional CTAB method was modified to extract high quality luffa DNA . Amplification for ISSR - PCR Test was approved.⑵The technical system of ISSR-PCR reaction for luffa was established and optimized. A 20μL reaction system, with 1.5 U TaqDNA polymerase , 0.4μmol / L primer, 250μmol / L dNTP, 30 ng template DNA and 2μL 10×Buffer was established. Under the system, 5 min pre-denaturation at 94℃, 30 s denaturation at 94℃, 45 s annealing at optimal annealing temperature, 90 s extension at 72℃, for 35 cycles, 10 min extension at 72℃after the end of the cycle, 4℃preserved. Amplified bands were clear, effective, and suitable for ISSR method analysis.⑶In the test 99 primers, 16 were selected for Luffa ISSR amplification ,which amplified high polymorphism, clear bands . Including 144 polymorphism bands ,166 bands were amplified .The polymorphism rate reached 86.75%. At the same time, the study found that the proportion of primers which contained (AG) n, (CA) n sequence were respectively up to 30%. illustrating the luffa genome rich in (TC)n and(GT)n microsatellite.⑷Acorrding to the ISSR analysis , the 30 varieties were classified by cluster analysis. The genetic similarity values were between 0.313 ~ 0.951, with an average of 0.769, illustrating a high genetic similarity. At GS = 0.313 luffa were devided into two types ,Luffa cylindrical (L.) Roem and Luffa (Luffa acutangula (L.) Roxb. revealling part of the genetic similarity general , genetic relationship and genetic distance of the thirty luffa varieties.⑸Through the Cluster analysis in the test (as shown in Figure 3-15), number 15, 9,17and 19 varieties had relatively greater genetic specificity.They could be used as parents to hybrid with other further relatives. Luffa cluster map revealed genetic similarity between varieties, and more for the future purpose of selective introduction, provided references for selected breeding and shortening the breeding period.