Genetic Diversity And Phylogenetic Relationship Of Impatiens In Ya’an Based On SRAP

Impatiens L. which has high ornamental value is one of the largest genus in extant angiosperm. With difficult specimen making and extremely obvious regionality and endemism in geographical distribution, classic classification work of Impatiens is extremely hard. The predecessors have not got uniform conclusion on Impatiens classification from morphological characteristics, such as leaves arrangement, leaves basal or cauline, inflorescence pattern, inflorescence flower number, flower detail feature, fruit and seedling morphology.The test select Ya’an area as the resource investigation and gathering places of wild plants in Impatiens. From2009to2011, based on field investigation at Daxing Town, Zhougong Mountain, Haizi Mountain, Bifeng Gorge, Horn River, Erlang Mountain, Taihu Temple, Longcang Ravine Forest Park in Ya’an, tender leaves of20species plants in Impatiens were collected and brought back to the laboratory. And then their genetic diversity were analyzed by SRAP markers to compare the genetic differences, analyze interspecific phylogenetic relationship and make cluster analysis. The purpose was provided the basis for classification, germplasm resources protection, the introduction and cultivation of breeding.The results were as follows:(1)DNA extracted by improved CTAB had better quality to meet requirements of SRAP markers.(2)SRAP-PCR amplification procedures and system of Impatiens had been established by screening annealing temperature gradiently, adjusting PCR program and optimizing SRAP reaction system orthogonally. Amplification procedures:95℃for5min, followed by5cycles of95℃for lmin,35℃for lmin and72℃for lmin,35cycles of95℃for lmin,49℃for lmin and72℃for lmin, and was terminated with5min extension step at72癈, save at4℃. Reaction system:50ng/25μl DNA,2.0mmol/L Mg2+,0.25mmol/L dNTPs,0.20μmo/L primers,1.5U/25μl Tαq DNA polymera.(3)Sixteen pairs of primers with higher polymorphism, better stability were screened in72, the test amplified407DNA fragments and25.4per primers, the size of the bands distributed in the range of0.1to2.5Kb, the percentage of polymorphic loci was100%.(4)Genetic similarity analysis of20species in Impatiens shows that, Nei’s genetic similarity coefficient distributed in0.5430to0.9410, the mean was0.6334. The coefficient between white and purple Ⅰ. rostellata, Ⅰ. oxyαnthera and Ⅰ. alpicola was the largest, between Ⅰ. wallerana and Ⅰ. rostellat, Ⅰ. waller ana and Ⅰ. lemeei was the smallest.(5)Genetic relationship tree of20species in Impatiens was drew by UPGMA.20species can be divided into3classes (class A, class B, class C) when the threshold was0.615, class C can be divided into4groups at0.636. There was a strong correlation between clustering result and morphological similarity.(6)The conclusion was drew by comparing and analyzing classification results comprehensively based on morphology, pollen morphology, molecular marker. Firstly, the morphological and pollen morphological characteristics of Ⅰ. pritzelii, Ⅰ. omeiαnα, Ⅰ. wilsonii are very similar, and they clustered into one group in the molecular cluster diagram, Ⅰ.lienchuanensis and Ⅰ. replans were the same as the former. They were classified as a category, this had been proved by the data from morphology, micro morphology, molecular biology. Secondly, tricolpate pollen and morphological features (such as leaves set at stem apices and unbranched,4pieces lateral sepals), four-trench pollen and morphological features (such as leaves powdered on stems,2pieces lateral sepals) both showed a strong correlation. Thirdly, pollen morphological characteristics (such as tricolpate, four-trench) and morphological characteristics (such as2or4pieces lateral sepals, leaves set at stem apices or not, peduncles or not and flower detail feature) can be used as the basis for classification in Impatiens.