Genetic Differentiation Of Micro-geographic Oligostachyum Scabriflorum Populations

Oligostachyum scabriflorum,a species of bamboo belonged to Oligostachyum. Oligostachyum scabriflorum had the biggest distribution area and reclaimed area among all species of this genus.The proliferation characteristic of Oligostachyum scabriflorum clone from Yong'an County was studied with inter-simple sequence repeat(ISSR)technique,and the genetic diversity and genetic structure of Oligostachyum scabriflorum were also discussed.The trait of genetic differentiation of Oligostachyum scabriflorum and its relationship with environmental factors were studied based on the former results.The results were as follows:(1)The reliable ISSR analysis system was established.In order to get the reliable system, the chemical components and the amplification program were optimized with grads selecting. The optimized ISSR-PCR conditions were as follows:20μl reaction volume containing 1.5 mmol/L Mg²⁺(1×Buffer with Mg²⁺),2 U Taq polymerase,0.8 mmol/L primer,0.1 mmol/L dNTPs, 50 ng DNA template.Furthermore,the addition of 1.5%formamide was propitious to reduce the disturbance.The optimized amplification program was:2 minutes at 94℃for predenaturing, then 45 cycles of 45 seconds at 94℃for denaturing,45 seconds at 52～58℃(different primers with different annealing temperatures)for annealing,2 minutes at 72℃for extending,finally extending at 72℃for 5 minutes after cycling,then stopped at 4℃.The PCR products were examined by agarose gel electrophoresis and the appropriate concentration of gel was 1.5%. 50 ISSR primers were used to screen the suitable primers for genetic assessing of Oligostachyum scabriflorum populations,of which 10 ISSR primers with high resolution and multiple polymorphic bands were screened.(2)Individuals of one clone had the same genetic structure,which clustered to one group based on Nei's genetic distance with ISSR technique.That was consistent with the structure of rhizome.12 Oligostachyum scabriflorum clones were distributed in the plot of 100 m² by clone judging with ISSR analysis.A plot Of 40 m×50 m was established and 30 samples of Oligostachyum scabriflorum individuals were collected at each intersection point on a 10 m×10 m grid within the plot.30 samples belonged to 9 clones,and each clone took up 2～5 intersection points meaning that the proliferation distance of Oligostachyum scabriflorum clones was 10～50 m.(3)The genetic differentiation index(Gst)ofOligostachyum scabriflorum population was 0.2470.The ratio of genetic variation caused by the different micro-geographic environment was 30%.The genetic clustering of Oligostachyum scabriflorum micro-geographic populations was inconsistent with geographic distribution.That was to say geographic separation was not the only reason for genetic differentiation.Oligostachyum scabriflorum flowered seldom,and it had few fruits after flowering.Its spread area was limited,and the gene flow was little,so the selection of environment played an important role in the genetic differentiation.The terrain and physiognomy was complex in Yong'an County,so the condition of water and heat changed greatly with the altitude changes,which lead genetic differentiation of the populations.(4)The relationships between genetic differentiation of Oligostachyum scabriflorum and the physical and chemical properties of soil were studied for the first time to find out the level of genetic diversity and the effect intensity of environmental factors on genetic differentiation.The results showed the negative correlations of genetic diversity indexes(including Na,N,H,I,PPB)of Oligostachyum scabriflorum with maximum water capacity,capillary water capacity, non-capillary porosity,capillary porosity,total porosity,soil air permeability in 0-20 cm and 20-40 cm soil layer,and positive correlation with soil density,all the correlations were not significant.The correlations of genetic diversity indexes were positive with pH value,total Nitrogen in 0-20 cm soil layer,but were negative with total Phosphorus,soil organic matter, C/N ratios.The correlations of genetic diversity indexes were positive with total Kalium except for Shannon diversity index,and the correlations were not significant.The correlations of genetic diversity indexes were positive with pH value,total Nitrogen,total Kalium,but were negative with total Phosphorus,soil organic matter,C/N ratios in 20-40 cm soil layer,and the correlations were not significant.The genetic differentiation of Oligostachyum scabriflorum was less depended on the physical and chemical properties of soil.We found that the relationships among genetic differentiation of Oligostachyum scabriflorum and altitude,slope position,longitude were positive,but were negative with slope degree and latitude,and the correlation levels were low.The difference of micro-geographic environmental factors caused the genetic differentiation of Oligostachyum scabriflorum,but the main factor was not clear.The environmental factors may influence combinatorially on genetic differentiation.