8. Determine reproducibility of DNA profiles by replicating all
RAPD reactions at least two times.
5.11 RAPD Data
Analysis
1. Use an average percentage similarity in band sharing to compare the degree of genetic variation between strains.
2. RAPD bands are scored for each primer as 1 for the presence
and 0 for the absence, and consistent RAPD bands are
considered.
3. Generate a similarity matrix for the analysis of individual primer
with each strain. These matrices are based on the percentage of
shared bands in ail pair-wise comparisons.
4. The equation F ¼ 2Cxy/(Tx + Ty) Â 100 (where F is the
similarity per primer, Cxy is the number of common bands,
and Tx and Ty are the total number of bands in that particular
pair-wise comparison) is used to generate a genetic similarity
matrix.
5. Use this matrix to perform hierarchical cluster analysis based on
the unweighted pair-group method algorithm (UPGMA).
6. Plot a dendrogram.
7. Use average linkage method [177] for cluster analysis on the
values of 1 À F, using the unweighted pair-group method
algorithm (UPGMA).
8. Plot dendrograms [178].
9. Subject the average percent similarities from each matrix to a
T test to determine if differences in genetic variation between
the strains are statistically significant.
10. The analysis is paired according to primers so that average
variation between strains is compared for each primer and
variation among primers is not a factor.
6 Phylogenetic Analysis
A phylogenetic structure is a vital component of the comparative
method in evolutionary biology and delivers a critical component
for studies of gene flow, population structure, biogeography,
coevolution, coadaptation, cospeciation, and historical ecology.
Systematic information is also important to rational implementation and monitoring agendas when these nematodes are preferred
as biological control agents [5]. The DNA sequences of the internally transcribed spacer 1 (ITS1) region of the ribosomal RNA
(rRNA) tandem repeat could provide the heritable characters requisite for a thorough phylogenetic analysis [172].
6.1 DNA Cloning
and Sequencing
1. Clone the purified ITS and 16S rDNA PCR product for each
strain into pGEM-T Easy vector.
Phylogenetic Analysis
93
RAPD reactions at least two times.
5.11 RAPD Data
Analysis
1. Use an average percentage similarity in band sharing to compare the degree of genetic variation between strains.
2. RAPD bands are scored for each primer as 1 for the presence
and 0 for the absence, and consistent RAPD bands are
considered.
3. Generate a similarity matrix for the analysis of individual primer
with each strain. These matrices are based on the percentage of
shared bands in ail pair-wise comparisons.
4. The equation F ¼ 2Cxy/(Tx + Ty) Â 100 (where F is the
similarity per primer, Cxy is the number of common bands,
and Tx and Ty are the total number of bands in that particular
pair-wise comparison) is used to generate a genetic similarity
matrix.
5. Use this matrix to perform hierarchical cluster analysis based on
the unweighted pair-group method algorithm (UPGMA).
6. Plot a dendrogram.
7. Use average linkage method [177] for cluster analysis on the
values of 1 À F, using the unweighted pair-group method
algorithm (UPGMA).
8. Plot dendrograms [178].
9. Subject the average percent similarities from each matrix to a
T test to determine if differences in genetic variation between
the strains are statistically significant.
10. The analysis is paired according to primers so that average
variation between strains is compared for each primer and
variation among primers is not a factor.
6 Phylogenetic Analysis
A phylogenetic structure is a vital component of the comparative
method in evolutionary biology and delivers a critical component
for studies of gene flow, population structure, biogeography,
coevolution, coadaptation, cospeciation, and historical ecology.
Systematic information is also important to rational implementation and monitoring agendas when these nematodes are preferred
as biological control agents [5]. The DNA sequences of the internally transcribed spacer 1 (ITS1) region of the ribosomal RNA
(rRNA) tandem repeat could provide the heritable characters requisite for a thorough phylogenetic analysis [172].
6.1 DNA Cloning
and Sequencing
1. Clone the purified ITS and 16S rDNA PCR product for each
strain into pGEM-T Easy vector.
Phylogenetic Analysis
93