Marine Invertebrates                    Dirk Steinke   Short Course on DNA Barcoding Methods   29 Nov 2011
Preservation                                       The good                                       • Ethanol preserved, not...
SamplingmtDNA rich tissue where possible• muscle tissue (larger animals e.g. fish)• legs (arthropods)• tube feet (echinode...
ExtractionRegular kits will do in many cases, butEchinodermsPolychaetesMollusks                    work much better with C...
PCRTaxon-specific primers are key    Check out http://connect.barcodeoflife.net/group/marinebarcoding
PCR   Majority works with                            Taxon specific primersone primer pair or cocktail                    ...
SequencingMultiplex primer cocktails      5’                                        3’                                    ...
Editing• some groups exhibit indels more frequently (e.g. crustacea, mollusks)• watch out for pseudogenes (often easy to s...
Thank you!
Upcoming SlideShare
Loading in …5
×

Dirk Steinke - Marine invertebrates

1,026 views

Published on

A short course on DNA barcoding methods for marine invertebrates.

Published in: Education, Technology
0 Comments
0 Likes
Statistics
Notes
  • Be the first to comment

  • Be the first to like this

No Downloads
Views
Total views
1,026
On SlideShare
0
From Embeds
0
Number of Embeds
3
Actions
Shares
0
Downloads
14
Comments
0
Likes
0
Embeds 0
No embeds

No notes for slide

Dirk Steinke - Marine invertebrates

  1. 1. Marine Invertebrates Dirk Steinke Short Course on DNA Barcoding Methods 29 Nov 2011
  2. 2. Preservation The good • Ethanol preserved, not older than 5 years • frozen tissue • RNA laterThe bad• Ethanol preserved, older than 10 years at room temp• DMSO (cross-reacts with Ethanol)• IsopropanolThe ugly• formalin• spirit•…
  3. 3. SamplingmtDNA rich tissue where possible• muscle tissue (larger animals e.g. fish)• legs (arthropods)• tube feet (echinoderms) The smaller the better!
  4. 4. ExtractionRegular kits will do in many cases, butEchinodermsPolychaetesMollusks work much better with CTAB extraction protocols (binds to polysaccharides that can inhibit PCR)
  5. 5. PCRTaxon-specific primers are key Check out http://connect.barcodeoflife.net/group/marinebarcoding
  6. 6. PCR Majority works with Taxon specific primersone primer pair or cocktail needed - Fish - Crustacea - Cephalopoda1 - Annelida - Gastropoda - Cnidaria - Pycnogonida - Porifera - Echinodermata - Brachipoda Notoriously difficult - Foraminifera - Bivalvia2 - Tunicata 1some families have a tandem copy of COI 2two different COI-versions (male/female)
  7. 7. SequencingMultiplex primer cocktails 5’ 3’ M13-tail • M13 tail for sequencing multiplex products • M13 also useful for standard primer pairs (Folmer-tailed version very successful)
  8. 8. Editing• some groups exhibit indels more frequently (e.g. crustacea, mollusks)• watch out for pseudogenes (often easy to spot through stop codons)• proper alignment is crucial• some symbiotic bacteria can be amplified using universal primers
  9. 9. Thank you!

×