DNA PROFILING OF WEED PLANTS AROUND THE AREAS OF HYDERABAD

1. DNA PROFILING OF WEED PLANTS
AROUND THE AREAS OF
HYDERABAD
Presented By,
R. Siva Dharshini,
II MSc.,Zoology
Avinashilingam Institute for Homescience and Higher
Education for Women, Coimbatore, Tamilnadu.
Guide:- Dr. Sunil Kumar Verma.

2. INTRODUCTION
• DNA profiling was the one of the
advanced technique for the
characterization and
authentication of individuals of
species.
• It will be useful for the
documentation species population
of the country.
• DNA profiling defines that
categorization of individual species
by the specific DNA markers.

3. • Identification of plant
species is of critical
importance in
conserving and utilising
biodiversity, but this
may be hindered by a
lack of taxonomic
expertise.
• Several DNA markers
are designed in SKV lab
for the DNA barcoding
of plants like mat K and
rbcL.

4. • Plant genome contains 3 billion base pairs which
are arranged in a particular sequence that gives
us a unique identity. But still the DNA is every
plant is almost similar. 90% of the DNA is same in
every plants (about 99.9% nucleotide bases are
exactly same in plants).
• DNA fingerprinting is based upon the rest 10%
difference in the plant DNA. This method is done
by matching the uncommon sequence of plants
with the suspect's unique sequence.
PRINCIPLE OF THE STUDY

5.  DNA ISOLATION
 AGAROSE GEL
ELECTROPHORESIS
 POLYMERASE CHAIN REACTION
 DNA SEQUENCING
 DNA SEQUENCE ANALYSIS
METHODS

6. DNA ISOLATION
Wash the leaves with water and Crush
them with liquid nitrogen
Incubate them with CTAB and 100µl
proteinase K for 3 to 4 hours at 560C
After incubation spin the samples at
10000rpm for 10mins and transfer the
supernatant to another eppendrof tube
Add equal amount of
chloroform:isoamylalcohol(24:1) and
mix it well and centrifuge at 10000rpm
for 10mins
Transfer the supernatant to another
tube and add 0.1volume of 7.5
ammonium acetate and mix well.
Add 1 volume of 100% ethanol and
Invert the tube slowly to precipitate
the DNA. Then place the tube at -20◦c
for 1-2 hours.
Remove the liquid by inverting the
tissue paper and add 300µl of 70 %
ethanol to wash the pellet.
Centrifuge at 12000 rpm for 3
minutes and discard the supernatant
slowly by using a tissue paper.
Air dry the pellet and add 200µl of
T.E buffer and store it the cold place
for further use.

7. AGAROSE GEL
ELECTROPHORESIS
• Agrose gel electrophoresis is used
to quantify the DNA.
• 0.8gm agrose with 1xTAE buffer
and microwave to melt it.
• Add 10µl of EtBr and make the gel
to solidify.
• Place the gel in gel box and load
the wells with loading dye, buffer
and DNA of samples and 1KB DNA
marker on one well.
• Run at 100V for 30mins and DNA
will separate according to the
quantity.
• We can see the bands through UV
illuminator and save the picture by
gel documentation system.

8. POLYMERASE CHAIN
REACTION
• To see the DNA
amplification, we use
PCR (Polymerase Chain
Reaction) technique.
• PCR was carried out by
using the Taq
polymerase.
• The universal primer for
matK DNA barcoding in
plants was used as a
primer for PCR.

9. DNA SEQUENCING & ANALYSIS
• By the amplification of DNA of the
sample, we have to give the exosap
treatment before the pcr products of
DNA to be sequenced.
• Then we can sequence the DNA by
the sanger DNA sequencing method.
• We have to analysis the DNA
sequences by the
‘AUTOASSEMBLER’ software with
the reference sequence.
• Analyzed sequences are used BLAST
to get score for identity, then
uploaded to the NCBI database for
authentication.

10. SUMMARY
• By the above methods, we get the identity of the
individual species.
• We were able to establish the identity of a total of
10 plants of the 26 weed plants used in this study.
The sequences from the remaining plants were
awaited, therefore could not be included in the
report.
• The sequences generated in this study will be
submitted in ncbi database.
• Further studies will continue in the laboratory of
SKV at CCMB.

11. SIGNIFICANCE
• DNA markers are helping hands for
forensics to identify the species history
within a minutes.
• DNA profiling will helpful to document
the biodiversity among species around
the world.
• The discovery of UNIVERSAL
PRIMER TECHNOLOGY by CCMB is
the big contribution to the world.
• It was useful to authenticate the species
to prevent illegal crime on endangered
species and wild medicinal plants and
their conservation.