Algae and algae products - Identification of the biomass of microalgae, macroalgae, cyanobacteria and Labyrinthulomycetes - Detection and identification with morphological and/or molecular methods

This document specifies a method for the detection and identification of microalgae, macroalgae, cyanobacteria and Labyrinthulomycetes by using morphological methods and/or molecular methods.
The morphological methods in this document are applicable to harvested wet biomass and to harvested dried unground biomass from microalgae, macroalgae, cyanobacteria and Labyrinthulomycetes that have been grown and/or harvested for further processing and/or use.
The molecular methods in this document are applicable to harvested wet biomass and to harvested dried and/or ground biomass from microalgae, macroalgae, cyanobacteria and Labyrinthulomycetes that have been grown and/or harvested for further processing and/or use.
This document describes a toolbox, consisting of several identification methods that can be chosen according to the applicability and purpose of the identification:
-   morphological methods based on observation and referring to scientific literature on taxonomy:
-   macroscopic identification;
-   light microscopic identification.
-   molecular methods for sequencing and blasting of sequences:
-   16S rDNA sequencing;
-   18S rDNA sequencing;
-   rbcL DNA sequencing;
-   ITS sequencing;
-   COX1 gene sequencing;
-   tufA gene sequencing.
This document does not deal with genetic purity of the biomass or quantification of the identified taxa.
This document is not suitable for the analysis of highly processed biomass with highly degraded DNA where the fragments’ length are not sufficient for amplification of the targets and the morphological characteristics cannot be assessed.

Algen und Algenprodukte - Identifizierung der Biomasse von Mikroalgen, Makroalgen, Cyanobakterien und Labyrinthulomycetes - Erkennung und Identifizierung mit morphologischen und/oder molekularen Verfahren

Dieses Dokument legt ein Verfahren für den Nachweis und die Identifizierung von Mikroalgen, Makroalgen, Cyanobakterien und Labyrinthulomycetes (Netzschleimpilze) mittels morphologischer und/oder moleku-larer Verfahren fest.
Die in diesem Dokument dargelegten morphologischen Verfahren sind anzuwenden für geerntete feuchte Biomasse und geerntete trockene, ungemahlene Biomasse von Mikroalgen, Makroalgen, Cyanobakterien und Labyrinthulo¬mycetes, die für die Weiterverarbeitung und/oder Verwendung kultiviert und/oder geerntet wurden.
Die in diesem Dokument dargelegten molekularen Verfahren sind anzuwenden für geerntete feuchte Biomasse und geerntete trockene und/oder gemahlene Biomasse von Mikroalgen, Makroalgen, Cyanobakterien und Labyrinthulomycetes, die für die Weiterverarbeitung und/oder Verwendung kultiviert und/oder geerntet wurden.
In diesem Dokument wird eine Sammlung von Verfahren beschrieben, die aus mehreren Identifizierungs-verfahren besteht, welche je nach Anwendbarkeit und Zweck der Identifizierung gewählt werden können:
-   morphologische Verfahren, die auf Beobachtung unter Bezug auf die wissenschaftliche Literatur zur Taxonomie basieren:
-   makroskopische Identifizierung;
-   lichtmikroskopische Identifizierung;
-   molekulare Verfahren zur Sequenzierung und zum Sequenzvergleich („Blasting“):
-   16S rDNS Sequenzierung;
-   18S rDNS Sequenzierung;
-   rbcL DNS Sequenzierung;
-   ITS Sequenzierung;
-   COX1 Gensequenzierung;
-   tufA Gensequenzierung.
Dieses Dokument behandelt weder die genetische Reinheit der untersuchten Biomasse noch die Quantifizierung der identifizierten Taxa.
Dieses Dokument ist nicht geeignet für die Analyse von stark verarbeiteter Biomasse mit hochgradig abgebauter DNS, bei der die Länge der Fragmente für die Amplifikation der Targets nicht ausreichend ist, und bei der die morphologischen Merkmale nicht beurteilt werden können.

Algues et produits d'algues - Identification de la biomasse de microalgues, macroalgues, cyanobactéries et Labyrinthulomycètes - Détection et identification à l'aide de méthodes morphologiques et/ou moléculaires

Le présent document spécifie une méthode de détection et d’identification des microalgues, macroalgues, cyanobactéries et Labyrinthulomycètes à l’aide de méthodes morphologiques et/ou moléculaires.
Les méthodes morphologiques utilisées dans le présent document sont applicables à la biomasse humide récoltée et à la biomasse sèche non broyée récoltée provenant des microalgues, macroalgues, cyanobactéries et Labyrinthulomycètes qui ont été cultivées et/ou récoltées en vue d’un traitement ultérieur et/ou d’une utilisation ultérieure.
Les méthodes moléculaires utilisées dans le présent document sont applicables à la biomasse humide récoltée et à la biomasse sèche et/ou broyée récoltée provenant des microalgues, macroalgues, cyanobactéries et Labyrinthulomycètes qui ont été cultivées et/ou récoltées en vue d’un traitement ultérieur et/ou d’une utilisation ultérieure.
Le présent document décrit une boîte à outils, comprenant plusieurs méthodes d’identification qui peuvent être choisies en fonction de l’applicabilité et de l’objectif de l’identification :
-   méthodes morphologiques reposant sur l’observation et faisant référence aux ouvrages scientifiques sur la taxonomie :
-   identification macroscopique ;
-   identification au microscope optique ;
-   méthodes moléculaires de séquençage et d’alignement de séquences :
-   séquençage de l’ADNr 16S ;
-   séquençage de l’ADNr 18S ;
-   séquençage de l’ADN rbcL ;
-   séquençage de l’ITS ;
-   séquençage du gène COX1 ;
-   séquençage du gène tufA.
Le présent document ne traite ni de la pureté génétique de la biomasse ni de la quantification des taxons identifiés.
Le présent document ne convient pas à l’analyse de la biomasse hautement traitée contenant de l’ADN fortement dégradé dans lequel la longueur des fragments n’est pas suffisante pour amplifier les cibles et dans lequel les caractéristiques morphologiques ne peuvent pas être évaluées.

Alge in izdelki iz alg - Ugotavljanje biomase pri mikroalgah, makroalgah, cianobakterijah in labirintulomicetah - Odkrivanje in prepoznavanje z morfološkimi in/ali molekulskimi metodami

General Information

Status
Published
Current Stage
6060 - Definitive text made available (DAV) - Publishing
Due Date
11-Aug-2021
Completion Date
11-Aug-2021

Buy Standard

Draft
prEN 17477:2020
English language
26 pages
sale 10% off
Preview
sale 10% off
Preview

e-Library read for
1 day

Standards Content (sample)

SLOVENSKI STANDARD
oSIST prEN 17477:2020
01-marec-2020
Alge in izdelki iz alg - Ugotavljanje biomase pri mikroalgah, makroalgah,

cianobakterijah in labirintulomicetah - Odkrivanje in prepoznavanje z morfološkimi

in/ali molekulskimi metodami

Algae and algae products - Identification of the biomass of microalgae, macroalgae,

cyanobacteria and Labyrithulomycetes - Detection and identification with morphological

and/or molecular methods

Algen und Algenprodukte - Identifizierung der Biomasse von Mikroalgen, Makroalgen,

Cyanobakterien und/oder Labyrinthulomycetes - Erkennung und Identifizierung mit
morphologischen und/oder molekularen Methoden

Algues et produits d'algues - Identification de la biomasse de microalgues, macroalgues,

cyanobactéries et/ou Labyrinthulomycètes - Détection et identification à l'aide de

méthodes morphologiques et/ou moléculaires
Ta slovenski standard je istoveten z: prEN 17477
ICS:
13.020.55 Biološki izdelki Biobased products
oSIST prEN 17477:2020 en,fr,de

2003-01.Slovenski inštitut za standardizacijo. Razmnoževanje celote ali delov tega standarda ni dovoljeno.

---------------------- Page: 1 ----------------------
oSIST prEN 17477:2020
---------------------- Page: 2 ----------------------
oSIST prEN 17477:2020
DRAFT
EUROPEAN STANDARD
prEN 17477
NORME EUROPÉENNE
EUROPÄISCHE NORM
January 2020
ICS 13.020.55
English Version
Algae and algae products - Identification of the biomass of
micro-algae, macro-algae, cyanobacteria and/or
Labyrithulomycetes - Detection and identification with
morphological and/or molecular methods
Algen und Algenprodukte - Identifizierung der
Biomasse von Mikroalgen, Makroalgen,
Cyanobakterien und/oder Labyrithulomycetes -
Erkennung und Identifizierung mit morphologischen
und/oder molekularen Methoden

This draft European Standard is submitted to CEN members for enquiry. It has been drawn up by the Technical Committee

CEN/TC 454.

If this draft becomes a European Standard, CEN members are bound to comply with the CEN/CENELEC Internal Regulations

which stipulate the conditions for giving this European Standard the status of a national standard without any alteration.

This draft European Standard was established by CEN in three official versions (English, French, German). A version in any other

language made by translation under the responsibility of a CEN member into its own language and notified to the CEN-CENELEC

Management Centre has the same status as the official versions.

CEN members are the national standards bodies of Austria, Belgium, Bulgaria, Croatia, Cyprus, Czech Republic, Denmark, Estonia,

Finland, France, Germany, Greece, Hungary, Iceland, Ireland, Italy, Latvia, Lithuania, Luxembourg, Malta, Netherlands, Norway,

Poland, Portugal, Republic of North Macedonia, Romania, Serbia, Slovakia, Slovenia, Spain, Sweden, Switzerland, Turkey and

United Kingdom.

Recipients of this draft are invited to submit, with their comments, notification of any relevant patent rights of which they are

aware and to provide supporting documentation.

Warning : This document is not a European Standard. It is distributed for review and comments. It is subject to change without

notice and shall not be referred to as a European Standard.
EUROPEAN COMMITTEE FOR STANDARDIZATION
COMITÉ EUROPÉEN DE NORMALISATION
EUROPÄISCHES KOMITEE FÜR NORMUNG
CEN-CENELEC Management Centre: Rue de la Science 23, B-1040 Brussels

© 2020 CEN All rights of exploitation in any form and by any means reserved Ref. No. prEN 17477:2020 E

worldwide for CEN national Members.
---------------------- Page: 3 ----------------------
oSIST prEN 17477:2020
prEN 17477:2020 (E)
Contents Page

European foreword ....................................................................................................................................................... 4

1 Scope .................................................................................................................................................................... 5

2 Normative references .................................................................................................................................... 5

3 Terms and definitions ................................................................................................................................... 6

4 Abbreviations ................................................................................................................................................... 8

5 Reagents ............................................................................................................................................................. 9

5.1 Reagents for morphological methods ...................................................................................................... 9

5.1.1 General ................................................................................................................................................................ 9

5.1.2 Tap water ........................................................................................................................................................... 9

5.1.3 Culture medium ............................................................................................................................................... 9

5.1.4 Phosphate-buffered saline (PBS) .............................................................................................................. 9

5.2 Reagents for molecular methods ............................................................................................................... 9

5.2.1 Thermostable DNA polymerase ................................................................................................................. 9

5.2.2 PCR reaction buffer ........................................................................................................................................ 9

5.2.3 Deoxynucleoside triphosphate mix (dNTPs) ........................................................................................ 9

5.2.4 Primer ................................................................................................................................................................. 9

5.2.5 Agarose gel ..................................................................................................................................................... 10

5.2.6 DNA Ladder .................................................................................................................................................... 10

6 Apparatus ........................................................................................................................................................ 10

6.1 General ............................................................................................................................................................. 10

6.2 Apparatus for morphological identification methods .................................................................... 10

6.2.1 Low-magnifying optical system, such as a magnifying glass or a dissecting

microscope ..................................................................................................................................................... 10

6.2.2 Light microscope .......................................................................................................................................... 10

6.2.3 Scientific literature, on taxonomy .......................................................................................................... 10

6.2.4 Microscopic slide .......................................................................................................................................... 10

6.2.5 Microscopic cover glass ............................................................................................................................. 11

6.3 Apparatus for molecular identification methods ............................................................................. 11

6.3.1 Thermocycler ................................................................................................................................................ 11

6.3.2 Gel electrophoresis device ........................................................................................................................ 11

6.3.3 DNA sequencer .............................................................................................................................................. 11

6.3.4 PCR tube .......................................................................................................................................................... 11

6.3.5 DNA free pipette ........................................................................................................................................... 11

6.3.6 Plugged sterile tip ........................................................................................................................................ 11

7 Principle .......................................................................................................................................................... 11

7.1 General ............................................................................................................................................................. 11

7.2 Morphological methods ............................................................................................................................. 11

7.3 Molecular methods ...................................................................................................................................... 12

8 Procedure........................................................................................................................................................ 12

8.1 General laboratory requirements .......................................................................................................... 12

8.2 Choice of the methods ................................................................................................................................ 12

9 Morphological identification methods ................................................................................................. 14

9.1 General ............................................................................................................................................................. 14

---------------------- Page: 4 ----------------------
oSIST prEN 17477:2020
prEN 17477:2020 (E)

9.2 Macroscopic observation with the naked eye or a magnifying glass ......................................... 14

9.3 Light microscopy ........................................................................................................................................... 14

9.3.1 General ............................................................................................................................................................. 14

9.3.2 Staining ............................................................................................................................................................. 14

9.3.3 Preparation of microscopic slides .......................................................................................................... 14

9.3.4 Microscopic observation ............................................................................................................................ 15

9.3.5 Identification using keys ............................................................................................................................ 15

10 Molecular identification methods .......................................................................................................... 15

10.1 DNA extraction ............................................................................................................................................... 15

10.2 PCR Amplification ......................................................................................................................................... 15

10.3 Primers ............................................................................................................................................................. 16

10.4 Control reaction ............................................................................................................................................ 16

10.5 Evaluation of PCR products ....................................................................................................................... 17

10.6 PCR product cloning .................................................................................................................................... 17

10.7 PCR Product sequencing ............................................................................................................................ 17

10.8 Evaluation of sequence data ..................................................................................................................... 18

10.9 Sequence analysis/comparison with reference sequences in public databases ................... 18

11 Test report ...................................................................................................................................................... 18

Annex A (informative) Examples of universally available primers........................................................ 20

A.1 Prokaryotic primers specific to cyanobacteria ................................................................................. 20

A.1.1 PCR _16S rDNA amplification ................................................................................................................... 20

A.1.2 Sequencing ...................................................................................................................................................... 20

A.2 Eukaryotic primers more general (microalgae; Labirynthulids; seaweeds) .......................... 20

A.2.1 PCR _18S rDNA amplification ................................................................................................................... 20

A.2.2 PCR primers for COX1 gene ....................................................................................................................... 20

A.2.3 PCR primers for tufA gene ......................................................................................................................... 21

A.2.4 PCR primers for rbcL ................................................................................................................................... 21

Annex B (informative) Scientific literature that may be used for identification ................................ 22

Annex C (informative) Examples of DNA extraction methods for PCR .................................................. 24

Annex D (informative) Example of a standard polymerase chain reaction (PCR) setup ................ 25

Bibliography ................................................................................................................................................................. 26

---------------------- Page: 5 ----------------------
oSIST prEN 17477:2020
prEN 17477:2020 (E)
European foreword

This document (prEN 17477:2020) has been prepared by Technical Committee CEN/TC 454 “Algae and

algae products”, the secretariat of which is held by NEN.
This document is currently submitted to the CEN Enquiry.

This document has been prepared under a mandate given to CEN by the European Commission and the

European Free Trade Association.
---------------------- Page: 6 ----------------------
oSIST prEN 17477:2020
prEN 17477:2020 (E)
1 Scope

This document specifies a method for the detection and identification of microalgae, macroalgae

(seaweed), cyanobacteria and Labyrinthulomycetes by using morphological methods and/or molecular

methods.

The morphological methods in this document are applicable to harvested wet biomass and to harvested

dried unground biomass from microalgae, macroalgae, cyanobacteria and Labyrinthulomycetes that

have been grown and/or harvested for further processing and/or use.

The molecular methods in this document are applicable to harvested wet biomass and to harvested

dried and/or ground biomass from microalgae, macroalgae, cyanobacteria and Labyrinthulomycetes

that have been grown and/or harvested for further processing and/or use.

This document describes a toolbox, consisting of several identification methods that can be chosen

according to the applicability and purpose of the identification:

— morphological methods based on observation and referring to scientific literature on taxonomy:

— macroscopic observation;
— light microscopic observation;
— molecular methods of sequencing and blasting of sequences:
— 16S-rDNA sequencing;
— 18S-rDNA sequencing;
— rbcL DNA sequencing;
— ITS sequencing;
— COX 1 gene sequencing;
— tufA gene sequencing.

This document does not deal with genetic purity of the biomass or quantification of the identified taxa.

2 Normative references

The following documents are referred to in the text in such a way that some or all of their content

constitutes requirements of this document. For dated references, only the edition cited applies. For

undated references, the latest edition of the referenced document (including any amendments) applies.

ISO 24276:2006, Foodstuffs — Methods of analysis for the detection of genetically modified organisms and

derived products — General requirements and definitions

FprEN 17399:2019, Algae and algae-based products or intermediates - Terms and definitions

---------------------- Page: 7 ----------------------
oSIST prEN 17477:2020
prEN 17477:2020 (E)
3 Terms and definitions

For the purposes of this document, the terms and definitions in FprEN 17399:2019, ISO 24276 and the

following apply.

ISO and IEC maintain terminological databases for use in standardization at the following addresses:

— IEC Electropedia: available at http://www.electropedia.org/
— ISO Online browsing platform: available at http://www.iso.org/obp
3.1
16S-rDNA sequencing

process of determining the sequence of nucleotides in a complete or partial gene coding for the 16S-

ribosomal ribonucleic acid

Note to entry 1: The largest amount of 16S rDNA gene sequencing work concerns prokaryotes.

Note to entry 2: DNA is deoxyribonucleic acid.
3.2
18S-rDNA sequencing

process of determining the sequence of nucleotides in a complete or partial gene coding for the 18S-

ribosomal ribonucleic acid

Note to entry 1: The largest amount of 18S rDNA gene sequencing work concerns eukaryotes.

Note to entry 2: DNA is deoxyribonucleic acid.
3.3
alignment

process or result of matching up the nucleotide residues of two or more biological sequences to achieve

maximal levels of identity
3.4
BLAST

sequence comparison algorithm optimized for speed used to search sequence databases for optimal

local alignments to a query

Note to entry 1: It directly approximates alignments that optimize a measure of local similarity, the maximum

signal pair (MST) score.
3.5
blasting of sequences
sequence comparison against commonly used gene sequence databases
3.7
COX1 gene sequencing
process of determining the sequence of nucleotides in the COX1 gene
3.8
detection
discovering the target organism/microorganism using a special method
---------------------- Page: 8 ----------------------
oSIST prEN 17477:2020
prEN 17477:2020 (E)
3.9
DNA extraction
separation of DNA from the other components in a test sample
Note to entry 1: Adapted from ISO 24276:2006
3.10
DNA sequence
order of nucleotides within a deoxyribonucleic acid molecule
3.12
GenBank
comprehensive public database of genetic reference sequences

Note 1 to entry: GenBank is part of the International Nucleotide Sequence Database Collaboration, which

comprises the DNA DataBank of Japan (DDBJ), the European Nucleotide Archive (ENA), and GenBank at National

Center for Biotechnology Information (NCBI). These three organizations exchange data on a daily basis.

3.13
ITS sequencing

process of determining the sequence of nucleotides in the internal transcribed spacer (ITS)

3.14
macroscopic observation
identification with the naked eye, based on taxonomic keys
3.15
microscopic observation

identification with magnification by using magnifying glasses, binoculars or microscopes, based on

taxonomic keys
3.16
molecular identification method

set of tools that rely on the comparison of the nucleic acid sequences of DNA obtained from an

organism/microorganism using the PCR (polymerase chain reaction)-based method with

public/documented data of known (micro)organisms

Note to entry 1: The data obtained using the respective follow-up tools like gene sequencing can be compared

with sequences of known species accessible via public databases (see 3.4).

Note to entry 2: These methods allow detection of low concentrations of DNA, non-viable organisms.

3.17
morphological identification method
identification method based on morphological characteristics
3.18
positive PCR control

known positive (identified) sample representating the DNA-sequence of the (micro)organism under

study

Note to entry 1: This control is used to demonstrate that the PCR reagents are working as intended.

Note to entry 2: Adapted from ISO 24276:2006
---------------------- Page: 9 ----------------------
oSIST prEN 17477:2020
prEN 17477:2020 (E)
3.19
rbcL DNA sequencing

process of determining the sequence of nucleotides (A, T, C, and G) in the gene that codes for the large

subunit of the protein ribulose-1,5-bisphosphate carboxylase/oxygenase
3.20
Sanger sequencing
cycle sequencing method using fluorescent-labelled dideoxynucleotides
3.21
strain identification
determination of the genus and preferably also of the species name of a strain
3.23
tufA gene sequencing
process of determining the sequence of nucleotides in the tufA gene
4 Abbreviations
A Adenosine
BLAST Basic Local Alignment Search Tool
C Cytosine
COX1 Cytochrome c oxidase subunit 1
DDBJ DNA Data Bank of Japan
DNA Deoxyribonucleic acid
ENA European Nucleotide Archive
G Guanine
GenBank GenBank at National Centre for Biotechnology Information (NCBI)
ITS Internal transcribed spacer
NCBI National Centre for Biotechnology Information
PCR Polymerase chain reaction
rbcL Ribulose-1,5-bisphosphate carboxylase/oxygenase
rDNA Ribosomal DNA
RNA Ribosomal ribonucleic acid
T Thymine
tufA Elongation factor TU
---------------------- Page: 10 ----------------------
oSIST prEN 17477:2020
prEN 17477:2020 (E)
5 Reagents
5.1 Reagents for morphological methods
5.1.1 General

Either of the following reagents shall be used when examining biomass for morphological identification

ensuring a certain amount of salts in the reagent, so that when examining the sample, bursting or

shrinking of cells is prevented. Distilled water shall not be used.
5.1.2 Tap water

Normal tap water usually contains sufficient amount of salts to prevent bursting of cells and can be used

to prepare a sample for microscopic observation.
5.1.3 Culture medium

Culture medium is used for culturing the organism to be identified contains sufficient amount of salts to

prevent bursting of cells and can be used to prepare a sample for microscopic oberservation.

5.1.4 Phosphate-buffered saline (PBS)

PBS is a commercially available water-based salt solution and it contains sufficient amount of salts to

prevent bursting of cells and thus, can be used to prepare a sample for microscopic oberservation

5.2 Reagents for molecular methods
5.2.1 Thermostable DNA polymerase

Thermostable DNA polymerase is a critical player in replicating the target DNA. It is an enzyme that is

derived from a thermophilic bacterium and functions at high temperature. It is used in the polymerase

chain reaction. Thermostable DNA polymerases are commercially available and should be used as

directed by the manufacturer’s protocol.
5.2.2 PCR reaction buffer

PCR reaction buffers are generally sold with the thermostable DNA polymerase (5.2.1). PCR reaction

buffers can include MgCl or the can come with a separate MgCl solution.
2 2
5.2.3 Deoxynucleoside triphosphate mix (dNTPs)

A deoxynucleoside triphosphate mix (dNTPs) consists of four basic nucleotides—dATP, dCTP, dGTP,

and dTTP. They are the building blocks of new DNA strands. These four nucleotides are typically added

to the PCR reaction in equimolar amounts for optimal base incorporation. dNTPs are commercially

available.
5.2.4 Primer

A primer is a synthetic DNA oligonucleotide of approximately 15–30 bases. They are designed to bind

(via sequence complementarity) to sequences that flank the region of interest in the template DNA.

Primers are used for PCR and sequencing reactions. For example, during PCR reaction, DNA polymerase

extends the primers from their 3′ ends. As such, the primers’ binding sites shall be unique to the vicinity

of the target with minimal homology to other sequences of the input DNA to ensure specific amplication

of the intended target.
See Annex A for PCR and sequencing primers recommendations.
---------------------- Page: 11 ----------------------
oSIST prEN 17477:2020
prEN 17477:2020 (E)
5.2.5 Agarose gel
Use an agarose gel of suitable concentration (e.g. 1 % to 2 % (w/v))
5.2.6 DNA Ladder

The DNA ladder is the molecular weight size marker. It is a set of standards that are used to identify the

approximate size of a molecule such as PCR product run on a gel during electrophoresis. DNA ladders

are commercially available.
6 Apparatus
6.1 General

The laboratory shall use properly maintained equipment according to the manufacturer's instructions

and should use the requirements given in ISO/IEC 17025. In addition to standard laboratory

equipment, specific apparatus is described in the individual standards.

Where available, calibration should be routinely performed on equipment where performance could

impact the data produced. Apart from the usual equipement, the following equipment is required.

6.2 Apparatus for morphological identification methods

6.2.1 Low-magnifying optical system, such as a magnifying glass or a dissecting microscope

For the morphological investigation of details, which are necessary for the identification of macroalgae

or tissues, a commercially available magnifying glass or a dissecting microscope might be sufficient.

These have lenses with magnifying factors between 0.65x and 5x. In the case of a dissecting microscope

complemented by the magnifying factor of the eyepiece (usually 10x) or lens in the light path of the

camera (often 10x as well). This results in a total magnification between 6.5x and 50x. Use as directed in

the apparatus’ manual.
6.2.2 Light microscope

For the morphological investigation of details of microscopic organisms, a commercially available light

microscope, optionally equipped with different contrasting and/or fluorescence units can be used.

These usually have front lenses with magnifying factors between 4x and 100x complemented by the

magnifying factor of the eyepiece (usually 10x) or lens in the light path of the camera (often 10x as

well). This results in a total magnification between 40x and 1000x. Optional contrasting equipment

such as phase contrast or Differential Interference Contrast (DIC, also called Nomarski contrast) or also

dark field mode might be useful depending on the taxonomic group to be studied. Use as directed in the

apparatus’ manual.
6.2.3 Scientific literature, on taxonomy

Choose scientific literature according to the organism to be identified. A selection is given in Annex B.

This literature might carry a dichotomous or polytomous key to guide you through the identification

process. Additional drawing or photos could help in identifying the correct taxon.

6.2.4 Microscopic slide

Microscopic slides are composed of glass and are commercially available in the typical size of

76 × 26 mm (length x width) and a thickeness of 1 mm to accommodate the x/y-table of a microscope.

Glass quality is optimized not to interfere with the optical light path and the front lenses. Before use, the

microscope slide should be cleaned so that no particles can interfere with the microscope details of the

organism to be investigated.
---------------------- Page: 12 ----------------------
oSIST prEN 17477:2020
prEN 17477:2020 (E)
6.2.5 Microscopic cover glass

Microscopic cover glasses are composed of glass and are commercially available in various sizes and

forms (round, squre, rectangular). For broad applications, square cover glasses with a size of

18 × 18 mm and a thickeness of 0,13-0,16 mm are recommendable. The thickeness of the cover glass is

important if higher magnifications are applied, such when using lenses > 50x magnifying factor; theses

usually are immersion objectives usually used with immersion oil. The thickeness of cover glass is

optimized for the light path providing an optimal vision of the organism studied.

6.3 Apparatus for molecular identification methods
6.3.1 Thermocycler

Thermocycler is a device used to amplify DNA or RNA samples by the polymerase chain reaction. The

thermocycler is programmed to raise and lower the temperature of the samples in a holding block

allowing for denaturation and reannealing of samples with various reagents. Amplified genetic products

can be used in many downstream applications such as cloning, sequencing, expression analysis and

genotyping.
6.3.2 Gel electrophoresis devic
...

Questions, Comments and Discussion

Ask us and Technical Secretary will try to provide an answer. You can facilitate discussion about the standard in here.