| Dendrocalamopsis beecheyana var.pubescens and Dendrocalamus minor var.amoenus introduced from Fujian,Guangxi and Guangdong area and mixed planting with Casuarina equisetifolia in the coastal sandy in the southern region of Fujian Province with wide adaptability,fast breeding,and highly developed root system.The study of the genetic diversity of the germplasm resources of these material is the key point of the high yield cultivation of Dendrocalamopsis beecheyana var.pubescens and Dendrocalamus minor var.amoenus in the coastal sandy area.In this paper,IS SR molecular markers was used to assess the genetic diversity in 17 populations of Dendrocalamopsis beecheyana var.pubescens and 16 populations of Dendrocalamus minor var.amoenus.At the same time,the genetic relationship and genetic variation among germplasm resources was analyzed and the DNA finger prints were established.All of these provide theoretical foundation for the identification of provenance,seed selection and conservation strategy.The conclusions are as follows:1.Constructed the optimum IS SR reaction system and reaction program for Dendrocalamopsis beecheyana var.pubescens:IS SR reaction mixtures had a total volume of 20 μL,which contained 2.5 mmol/L Mg2+,0.25 mmol/L dNTPs,1.25 U Taq DNA polymerase,0.6μmol/L primer,10 ng template DNA,2 μL 10xBuffer,10.55 μL ddH2O.reaction program:pre-denaturation at 94℃ for 5 min,denaturation at 94℃ for 1 min,annealing at 54.5℃ for 0.5 min,extension at 72℃ for 1.5 min,40 cycles,final extension at 72℃ for 10 min,remaining at 4℃.2.Constructed the optimum IS SR reaction system and reaction program for Dendrocalamus minor var.amoenus:ISSR reaction mixtures had a total volume of 20 μL,which contained 2.5 mmol/L Mg2+,0.25 mmol/L dNTPs,1.50 U Taq DNA polymerase,0.5 μmol/L primer,80 ng template DNA,2 μL 10xBuffer,9.5 μL ddH2O.reaction program:pre-denaturation at 94℃ for 5 min,denaturation at 94℃ for 1 min,annealing at 52.7℃ for 0.5 min,extension at 72℃ for 1.5 min,40 cycles,final extension at 72℃ for 10 min,remaining at 4℃.3.16 IS SR primers screened from 100 primers were selected out for the amplification of Dendrocalamopsis beecheyana var.pubescens.A total of 185 bands amplified by ISSR primers,in which 167 bands were polymorphic bands,the percentage of polymorphic band 90.27%;12 ISSR primers were selected out for the amplification of Dendrocalamus minor var.amoenus.A total of 124 bands amplified by ISSR primers,in which 102 bands were polymorphic bands,the percentage of polymorphic band 82.26%.4.The genetic diversity of 17 populations of Dendrocalamopsis beecheyana var.pubescens was as follows:Nei’s gene diversity(He)was 0.2497,Shannon’s information index(Ⅰ)was 0.3949.The highest genetic variability was present in the Western Fujian distribution:Nei’s gene diversity(He)was 0.2315,Shannon’s information index(Ⅰ)was 0.3358,and the lowest levels were present in the Eastern Guangxi distribution:Nei’s gene diversity(He)was 0.0673,Shannon’s information index(Ⅰ)was 0.0968.The within-distributions component accounted for 53.99%of the total variations,while the among-distributions component accounts for 46.11%.The level of gene flow(Nm)was calculated as 0.4261 individuals per generation within the six distributions,indicating that there was little gene flow between the distributions.5.The genetic diversity of 16 populations of Dendrocalamus minor var.amoenus was as follows:Nei’s gene diversity(He)was 0.0460,Shannon’s information index(Ⅰ)was 0.0659,The highest genetic variability was present in the Minhou,Fuzhou population:Nei’s gene diversity(He)was 0.1162,Shannon’s information index(Ⅰ)was 0.1664.The lowest levels were present in the Conghua,Guangzhou population:Nei’s gene diversity(He)was 0.0064,Shannon’s information index(Ⅰ)was 0.0093.At the distribution level,Nei’s gene diversity(He)was 0.2066,Shannon’s information index(Ⅰ)was 0.3005.The highest genetic variability was present in Fujian distribution:Nei’s gene diversity(He)was 0.2987,Shannon’s information index(Ⅰ)was 0.4388.The lowest levels were present in Guangdong distribution:Nei’s gene diversity(He)was 0.1493,Shannon’s information index(Ⅰ)was 0.2150.6.The highest D(0.8383)of Dendrocalamopsis beecheyana var.pubescens was found between the ZL and CTB,indicting a closer genetic relationship.The lowest D(0.0386)was found between the QGG and CTB,indicting a farthest genetic relationship.At the distribution level,The lowest D(0.0564)was found between the Western Fujian and Central Fujian distributions,which indicated that these distributions was the most closely related.The highest D(0.4590)was found between the Southern Fujian and Eastern Guangxi distributions,indicating a wide gap in the relationship of these distributions.The D obtained from our IS SR analysis demonstrated a low genetic diversity among Dendrocalamopsis beecheyana var.pubescens distributions.The 17 populations were found to form six major groups and 6 distributions were clustered into three groups in an UPGMA dendrogram.7.The genetic distance between 16 populations of Dendrocalamus minor var.amoenus was 0.0139-0.8176 range of genetic distance.The populations of genetic relationship between TD and YQ were closer,the populations of genetic relationship between YS and WL were farthest.At the distribution level,genetic relationship between Guangdong and Guangxi were closer and the genetic distance was 0.0128,genetic relationship between Guangxi and Fujian was farthest and the genetic distance was 0.1003.The D obtained from our ISSR analysis demonstrated a low genetic diversity among Dendrocalamus minor var.amoenus distributions.The 16 populations were found to form three major groups in an UPGMA dendrogram.The clustering in this dendrogram was not consistent with the geographical distribution,suggesting that there was no obvious correlation between genetic distance and geographic distance of populations.8.According to the ISSR results,the DNA fingerprints of the series 17 populations of Dendrocalamopsis beecheyana var.pubescens and 16 populations of Dendrocalamus minor var.amoenus were established.The materials can be identified effectively based on the bands’ occurs,mount,and the styles of patterns amplified with different combinated primers. |
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