Name:

Lab Section: Unknown assigned:

Items to include in your Canvas Lab Notebook - to be submitted AS INDIVIDUALS!

1. Title and Date: An informative project title. Also, give the date your DNA Subway analysis
was completed.
1. DNA Sequencing to Determine an Unknown Specimen from the JMU Arboretum
2. Mar. 30, 2021

2. DNA subway Blue Line Project Info: Your DNA subway login and blue line project title in
the format provided at the beginning of the lab handout. Also include your project ID#.
1. laythenm
2. Laythe_plant_30Mar2021_BIO140
3. 230034

3. Consensus sequence: A screenshot of your consensus sequence. Be sure the image

includes the length of the consensus sequence. If you only have one sequence, include a
screenshot of the length of that sequence.

4. BLAST hits: A screenshot of your top seven to ten BLAST hits. This should include all
columns (alignment length, bit score, e-value, and mismatches) for each hit. Also make sure
you include the column headers in your screenshot.
 5. Taxonomic Table: Fill in the table with information about the top three taxonomically
unique hits in your BLAST search. You can find the required information by either clicking
on the links in your BLAST results or doing a google search for each hit and checking
Wikipedia (taxonomic information is listed on the right).

Taxonomic info Alignment length Bit score e-value mismatches

K: Plantae

P: Tracheophytes

C: Monocotyledon
562 1
O: Asparagales
993 0.0
F: Amaryllidaceae

G: Narcissus

S: N. tazetta

O: Asparagales 571
F: Amaryllidaceae
991 0.0 5
G: Pancratium

S: P. canariense

O: Asparagales 0.0
F: Amaryllidaceae
570 989 5
G: Pancratium

S: P. maritimum

K=Kingdom, P=Phylum, C=Class, O=Order, F=Family, G=Genus, S=Species

* assume K, P, and C are the same as the first table entry

6. BLAST Best match: Give the taxonomic information for the BLAST hit that is the BEST
match to your specimen. Explain how you know this is the best match. Your answer must
be in a complete sentence(s) and include discussion of alignment lengths,
mismatches and bit scores.

Taxonomic Information: Narcissus Tazetta subsp. chinensis

How do you know it is This is the best match as it has the highest bit
the best match? score over the matches. Narcissus Tazetta
has a similar alignment length to the
unknown. Lastly, as it only has 1 mismatch,
Narcissus Tazetta has less sequence
differences with the unknown than other
matches, indicating it is likely to be the most
closely related.

7. MUSCLE alignment: A screenshot of your trimmed MUSCLE alignment. 

8. Sequence similarity matrix: A screenshot of your sequence similarity matrix.

9. Most similar MUSCLE match: Which BLAST hit showed the highest percent similarity to
your sample after your MUSCLE alignment? What % similarity did it have?

Blast Hit: 3: Narcissus_tazetta

% Similarity 99.82
10. Least similar MUSCLE match: Which sample showed the lowest percent similarity to your
sample after your MUSCLE alignment? What % similarity did it have?  Make sure you
understand why you should use this as the outgroup for your trees.

Blast Hit: 1: Allium_altaicum-Allioideae

% Similarity 95.89

11. NJ tree: A screenshot of your NJ tree with the properly selected outgroup and YOUR
SAMPLE HIGHLIGHTED. Based on only the NJ tree, name the most specific taxonomic
group (e.g. Canis) to which you are confident your specimen belongs. (You must list
taxonomic information for taxa on the tree not in the top 3 Blast hits if it is needed to name
the most specific group you are confident your specimen belongs to.)

The most specific taxonomic group to which the Unknown specimen, according to the NJ
tree, is the Family, Amaryllidaceae.

Taxonomic info for Lycoris Squamigera:

O: Asparagales
F: Amaryllidaceae
G: Lycoris
S: L. Squagmigera

12. ML tree: A screenshot of your ML tree with the properly selected outgroup and YOUR
SAMPLE HIGHLIGHTED. Based on only the ML tree, name the most specific taxonomic
group (e.g. Canis) to which you are confident your specimen belongs. (You must list
taxonomic information for taxa on the tree not in the top 3 Blast hits if it is needed to name
the most specific group you are confident your specimen belongs to.)
 The most specific taxonomic group to which the Unknown specimen, according to the ML
tree, is the Family, Amaryllidaceae.

13. Google/Wikipedia images of top three BLAST hits: Include at least one photo of YOUR
organism. For students working with the unknown samples, image files are included in your
unknown folder. Also include Google/Wikipedia images of top three BLAST hits. You can do
less than three Google/Wikipedia images if there aren't three obviously different hits.
Compare your image(s) with the BLAST hit images to see which BLAST hit image your
unknown most closely resembles to prepare you to answer Question 14.

1. Unknown
2. Narcissus tazetta

3. Pancratium canariense
 4. Pancratium maritimum

5. Lycoris squamigera

14. Organism ID: Based on the results from your BLAST search (question 6 above), MUSCLE
alignment (question 9 above), phylogenetic trees (questions 11 and 12 above), and your
image search (question 13 above), fill in the table immediately below with the most specific
taxonomic level to which you are confident your specimen belongs. Include the taxonomic
level (Order, Family, Genus, Species, etc.), and the taxon name.

Taxonomic Level Family

Taxon name Amaryllidaceae

Fill in the table below with the higher level taxonomic information for the taxon you named
immediately above; only fill in rows down to and including the level to which you are
confident your unknown belongs.

Kingdom Plantae
Phylum Tracheophytes
Class Monocotyledon
Order Asparagales
Family Amaryllidaceae
Genus Unknown***
Species Unknown***