A protocol of polymerase chain reaction-random amplified polymorphic DNAs (PCR-RAPDs) was established to analyse the gene diversity and genotype identification for clones of Sequoia sempervirens (D. Don) Endl. in Chile. Ten (out of 34) clones from introduction trial located in Voipir-Villarrica, Chile, were studied. The PCR-RAPDs technique and a modified hexadecyltrimethylammonium bromide (CTAB) protocol were used for genomic DNA extraction. The PCR tests were carried out employing 10-mer random primers. The amplification products were detected by electrophoresis in agarose gels. Forty nine polymorphic bands were obtained with the selected primers (BG04, BF07, BF12, BF13, and BF14) and were ordered according to their molecular size. The genetic similarity between samples was calculated by the Jaccard index and a dendrogram was constructed using a cluster analysis of unweighted pair group method using arithmetic averages (UPGMA). Of the primers tested, 5 (out of 60) RAPD primers were selected for their reproducibility and high polymorphism. A total of 49 polymorphic RAPD bands were detected out of 252 bands. The genetic similarity analysis demonstrates an extensive genetic variability between the tested clones and the dendrogram depicts the genetic relationships among the clones, suggesting a geographic relationship. The results indicate that the RAPD markers permitted the identification of the assayed clones, although they are derived from the same geographic origin.
Genotype ; Polymerase Chain Reaction ; methods ; Random Amplified Polymorphic DNA Technique ; methods ; Sequoia ; classification ; genetics

OBJECTIVETo establish an effective way for rapid identification of Monascus strains based on DNA molecular marker.

METHODA random amplified polymorphic DNA (RAPD) marker named F421 in genomic DNA of Monascus F strain was observed during a comparison of DNA fingerprints derived from 10 cultivated strains of Monascus. F421 was cloned and sequenced. Comparing the sequence of F421 (GenBank accession number EF063107) with other relative sequences in the GenBank databases, no distinct comparability was found. A pair of sequence characterized amplified region (SCAR) primers were designed based on the sequence of the cloned fragment and tested for the specific detection of Monascus F.

RESULTThe results of polymerase chain reaction showed that only a 421bp segment of Monascus F strain was amplified compared with other 9 cultivated strains of Monascus. And the acquired SCAR marker of strain F could be used as a specific DNA fingerprint to identify Monascus strain F within one day.

CONCLUSIONSCAR molecular marker technology is an effective new way to identify Monascus strains more rapidly. And also is an assistant tool to identify Monascus strains more accurately when disagreements come out using traditional classification. It could be applied widely to the protection of germ plasm resources, classification and identification distinguishing false strains of pharmaceutical fungi.

Molecular Sequence Data ; Monascus ; classification ; genetics ; Polymerase Chain Reaction ; methods ; Random Amplified Polymorphic DNA Technique

OBJECTIVETo analyze the genetic diversity of wild Rehmannia glutinosa and evaluate and compare random amplified polymorphic DNA (RAPD) and inter sample sequence repeat (ISSR) for analysis of R. glutinosa accessions.

METHODTwo molecular markers, RAPD and ISSR were used for analyzing 55 wild R. glutinosa accessions.

RESULTAverage 16.00 and 19.08 bands were amplified by RAPD primers and ISSR primers respectively, and the percentage of polymorphic bands were 89.58% and 94.32% respectively; Fifty-five R. glutinosa accessions categorized into 7 clusters were identified by unweighted pair-group method, arithmetic average (UPGMA) method.

CONCLUSIONA high level of genetic diversity of wild Rehmannia glutinosa was displayed at DNA level, and genetic diversity coefficient of R. glutinosa from different production areas was 0.63-0.98, and ISSR marker can detect higher genetic diversity of R. glutinosa germplasms than RAPD marker.

Genetic Variation ; genetics ; Phylogeny ; Random Amplified Polymorphic DNA Technique ; methods ; Rehmannia ; classification ; genetics

OBJECTIVETo establish a new method for the identification of Schisandra sphenanthera and S. chinensis.

METHODRandom amplified polymorphic DNA-Sequence characterized applied region (RAPD-SCAR) method was applied to screen primers.

RESULTScreening from 100 primers, only 2 random primers, which can be used to identify S. sphenanthera and S. chinensis accurately with a good reproducibility. It worked to fit them into sequence characterized applied region.

CONCLUSIONRAPD-SCAR can be used to identify S. sphenanthera and S. chinensis accurately.

Base Sequence ; Molecular Sequence Data ; Polymerase Chain Reaction ; methods ; Random Amplified Polymorphic DNA Technique ; Schisandra ; genetics ; Sequence Analysis, DNA

Forensic botany is a study of judicial plant evidence. Recently, researches on DNA labeling technology have been a mainstream of forensic botany. The article systematically reviews various types of DNA labeling techniques in forensic botany with enumerated practical cases, as well as the potential forensic application of each individual technique. The advantages of the DNA labeling technology over traditional morphological taxonomic methods are also summarized.
Amplified Fragment Length Polymorphism Analysis ; Botany/methods* ; DNA Fingerprinting/methods* ; DNA, Plant/analysis* ; Forensic Genetics ; Minisatellite Repeats ; Random Amplified Polymorphic DNA Technique

BACKGROUNDPenicillium marneffei (P. marneffei) is an emerging pathogenic fungus that can cause invasive mycosis in patients with AIDS. The epidemiological features of P. marneffei infection in AIDS patients in Guangdong province remain unclear so far. This study aimed to investigate the genetic diversity within a population of 163 P. marneffei isolates obtained from AIDS patients and search for the dominant clinical strains in Guangdong province.

METHODSOne hundred and sixty-three P. marneffei isolates obtained from AIDS patients in Guangdong province during January 2004 and December 2009 were studied by randomly amplified polymorphic DNA (RAPD) using two random primers (H2 and H22). The degree of similarity between samples was calculated through similarity coefficients from RAPD fragment data and the dendrogram was assessed using the unweighted pair group method with arithmetic mean (UPGMA).

RESULTSTwo primers showed a high degree of discrimination and good stability. Primer H2 yielded eight different patterns (H2-1 to H2-8) among 163 isolates with the discriminatory power being 0.413. Primer H22 identified seven types (H22-1 to H22-7) among 163 isolates with the discriminatory power being 0.467. Genetic similarity coefficients based on RAPD data among 163 P. marneffei isolates ranged from 0.681 to 0.957, 61.96% of which were no less than 0.83. The discriminatory power of the two primers was 0.524. One hundred and sixty-three P. marneffei isolates were clustered into nine distinct groups (groups I to IX) at the similarity coefficient value of 0.83 and group I was the most common, including 101 strains (61.96%).

CONCLUSIONThe RAPD analyses could provide important information as to the degree of genetic diversity and the relationship among clinical P. marneffei isolates, revealing genetic polymorphism and dominant strains.

Acquired Immunodeficiency Syndrome ; microbiology ; Genetic Variation ; genetics ; Humans ; Penicillium ; classification ; genetics ; Random Amplified Polymorphic DNA Technique ; methods

In order to produce interspecific somatic hybrids between Brassica campestris (2n = 20, AA) and Brassica oleracea (2n = 18, CC), we isolated protoplasts from cotyledons and hypocotyls of young seedlings, and fused by 40% polyethylene glycol (PEG). Fused cells were cultured in modified K8p liquid medium supplemented with 0.2 mg/L 2,4-dichorophenoxyacetic acid (2,4-D) +0.5 mg/L 6-benzylaminopurine (6-BA)+0.1 mg/L naphthaleneacetic acid (NAA)+ 0.1 mg/L Kinetin (Kin), 0.3 mol/L sucrose and 0.3 mol/L glucose were used as osmoticum. At the eight-to ten-cell stage, divided cells were transferred to Kao's basal medium supplemented with 0.3 mol/L sucrose as carbon source and 0.1% agarose, 2 mg/L 6-BA+ 2 mg/L Zeatin (ZEA)+1 mg/L NAA+ 0.5 mg/L Kin for callus induction. After 35 days, when small calli reached 2-3 mm in diameter, calli were transferred to regeneration medium containing 5 mg/L Zeatin (ZEA) and 2 mg/L indole-3-acetic acid (IAA). After the length of the shoots reached 1-2 cm, the shoots were transferred to 1/2 MS+0.2 mg/L NAA for root induction. Morphological, cytological and molecular biological analysis methods were used for identification of somatic hybrids. The results showed that, the first cell division occurred during 2-7 days of culture. Five weeks after culture initiation, the plating efficiency attained 0.66%. Finally, the shoot regeneration frequency was 3.7%. A total of eleven regenerated plants were obtained and verified as somatic hybrids by morphological observation and flow cytometry. Cytological studies showed that all tested plants had a chromosome number of 38, the sum of both parents. Hybridity was also confirmed by randomly amplified polymorphic DNA (RAPD) and genomic in situ hybridization (GISH) analysis, indicating that these regenerated plants were all true hybrids of B. campestris and B. oleracea. All amphidiploid somatic hybrids showed low pollen fertility. Pollen fertility was gradually recovered in the first and second progenies.
Brassica ; genetics ; growth & development ; physiology ; Breeding ; methods ; Genes, Plant ; Hybridization, Genetic ; genetics ; Protoplasts ; physiology ; Random Amplified Polymorphic DNA Technique

OBJECTIVE: The feasibility of Papaver somniferum L. cultivars identification was explored by TD-RAPD technique. METHOD: Genomic DNA was extracted by improved CTAB method. One sample of species from Papaver somniferum L in xishuangbanna area. was studied by using TD-RAPD method. RESULT: We established an optimal method of extracting genomic DNA. Six primers were picked out from 10 primers. CONCLUSION TD-RAPD could be applied to researches of molecular marker of Papaver somniferum L. TD-RAPD technique provide a method to constitute DNA database of Papaver somniferum L. and conclude the source of opium poppy.
DNA Primers ; DNA, Plant/isolation & purification* ; Feasibility Studies ; Humans ; Papaver/genetics* ; Plant Leaves/genetics* ; Polymerase Chain Reaction ; Random Amplified Polymorphic DNA Technique/methods* ; Species Specificity

OBJECTIVETo provide reference for breeding of cultivated ginseng by researching the genetic diversities among different strains.

METHODTo make up the systematic diagram of genetic relationship by calculating the proximity coefficient with SPSS 10.0 software and clustered by between-groups linkage method.

RESULTThe selected 18 random primers used in PCR amplification to produce 145 bands in 17 samples, among which 53 bands (36.5%) are polymorphic. The results of cluster analysis show that the genetic relationship between different strains of Da-maya is closer than that of Er-maya, which proves that it is feasible to differentiate the cultivated groups in molecular level on outside character.

CONCLUSIONRAPD is a favorable molecular marker to assist ginseng breeding.

Cluster Analysis ; DNA Primers ; genetics ; DNA, Plant ; genetics ; Genetic Variation ; Panax ; classification ; genetics ; growth & development ; Phylogeny ; Plants, Medicinal ; genetics ; growth & development ; Random Amplified Polymorphic DNA Technique ; methods

In the present study, the genome shuffling was used to improve lipase production of Penicillium expansum. A lipase producing mutant strain-Penicillium expansum FS8486 and a wild type of Aspergillus Tamarii FS-132 isolated from soil of a volcano in Xinjiang were used as the parental strains. After two rounds of genome shuffling, several elite daughter strains were screened. The lipase activity in one of the daughter strains was increased 317% over the starting strain FS8486. Comparisons of the morphology, RAPD (Random Amplification of Polymorphic DNA) polymorphism and the fatty acid compositions between the daughter and the parental strains suggested that the filial generation were generated by genome shuffling. In this study, the genome shuffling used successfully first time in eukaryotic microorganism and increases the production of the desired metabolite in short time, the study will be useful to spread the genome shuffling in eukaryotic microbial breeding.
Aspergillus ; genetics ; DNA Shuffling ; methods ; Genetic Enhancement ; methods ; Genome, Fungal ; genetics ; Lipase ; biosynthesis ; genetics ; Penicillium ; enzymology ; genetics ; Random Amplified Polymorphic DNA Technique