field collection and sampling 1courtesy of carol ritland

Field Collection and Sampling

1courtesy of Carol Ritland

Starting your collection• What to sample: Tissue

• What to consider: Age, Season

• What methodology to used for your biological questions

• What is the natural

history of your organism

eg. Life history etc.http://images.google.ca/imgres?imgurl=http://www.christiananswers.net/kids/animals6.jpg&imgrefurl=http://www.christiananswers.net/kids/find2.html&h=375&w=500&sz=70&hl=en&start=5&usg=__d6sSz-0SC0vP_NuogTY5sRsvsxY=&tbnid=a9-2ftNtkcqeBM:&tbnh=98&tbnw=130&prev=/images%3Fq%3Danimals%26gbv%3D2%26hl%3Den%26client%3Dfirefox-a%26rls%3Dorg.mozilla:en-US:official%26sa%3DG

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Starting your collection….cont’d

• Check for permission and necessary permits for sampling

• Check for location of field sites

• Check on logistics of field collection

http://www.monkeygrove.com/scribbles/2002/PaperWork.gif3

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Conifer abundance: affect on soil richness

Photos = L. Ritlandcourtesy of Carol Ritland

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Starfish abundance: affect on barnacles

Photos = L. Ritland

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1) What should you collect?

2) What could you collect?

3) Where would you collect?

4) When would you collect?

5) How would you collect?

6) How much would you collect?

7) Why did you collect them?

Tissue (Always go fresh if possible):

– Plant: Seed, Leaves, Flower, Pollen, Bark, Xylem, Roots

–Animal: Reproductive tissue, muscle, skin, hair, scat, blood, ear/toe/tail clip, fin, tooth, sloughed skin, saliva

–Fungal: Hyphae, spores, fruiting bodies

–Bacterial: single isolate culture

–Destructive vs Non Destructive methods

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Factors to consider when collecting:

• Age:– Plant = actively growing material such as

apical points, seedlings– Animal = actively dividing tissue (buccal and

blood cells)– Fungus = young fruiting bodies (pure culture)– Bacteria = liquid culture

• Season:– For conifer = use early spring bud burst

especially for DNA markers8

How to sample:

• Sampling schemes:– Linear – Quadratic squares– Distance between samples – For animals: migration, reproductive

strategies, life cycle– For plants: clonality, roots, reproductive

strategies, life cycle– Consult a statistician? (Gene expression

studies)9courtesy of Carol Ritland

Sampling cont’d

• Tools for sampling:– Ideally flash freezing samples with liquid

nitrogen and transportation under ultra low temperature

– Clean and if possible sterilize collection tools between samples

– Pack samples with foil or proper containers eg. cryovials

– Label all samples with non water based ink and protect with clear tape/paper and pencil

10courtesy of Carol Ritland

Liquid nitrogen vapour tanks

11courtesy of Carol Ritland

Ultralow temp. Liq N2 tanks

•Pending on size•Ultralow temp can last from 10 days to 3 weeks•Can be used on aircrafts

12courtesy of Carol Ritland

Use sterile technique when

sampling

•Keep meticulous records•Identify any problems in the field for given sample•Do not depend on your memory alone

13courtesy of Carol Ritland

Long Term Storage:

– Ideally all tissues should be kept at ultra low temperature (minus 70 to 80°C)

– Certain tissues (blood, animal tissues cut into small pieces <1mm2 ) could be stored in 100% ethanol and saturated EDTA

– Plants (small amounts) could be desiccated with lots of silica beads with lots of changes of beads

14courtesy of Carol Ritland

No No..s

• Items to avoid:– Unnecessary chemicals eg. Formalin– historical samples that has been treated with

fixatives– degraded samples– freezing and thawing of tissue– freezer burn (improper storage conditions)– improper inventory of samples

15courtesy of Carol Ritland

More to ponder…..

• Items to consider:– For certain molecular marker (eg. AFLP,

microarray) use the same tissue type (developmental differences could cause error when genotyping)

– If possible collect all samples within a season over a same span of time (eg. For microarray analysis)

– Collect more samples than required for pilot study and lost of samples

16courtesy of Carol Ritland