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HHMIHoward Hughes Medical Institute

Tufts UniversityDepartment of Chemistry

Mitochondrial DNA: Where Chemistry, Biology and Anthropology Meet

Meredith Knight, Tufts UniversityRocco Cieri, Medford High School



Presentation Overview

• Concept• Science• Development • Experiment• Dissemination • Questions



Concept – “From spit to SNP”

• Develop an experiment to allow high school and college students to determine their “deep ancestry” using the mtDNA in their saliva.

AdvantagesEngaging for studentsReal world applicationGenetics

ChallengesKeeping cost lowLimited timeLimited equipment



Science Background

Mitochondria are in all cells

Mitochondria contain DNA

Wikipedia - File:Biological_cell.svg

Wikipedia - Animal_mitochondrion_diagram_en_(edit).svg‎



Science BackgroundMitochondrial DNA is only inherited from mother; does not recombine



Science Background – Mitochondrial DNA (mtDNA)

• Human mtDNA = 16,569 base pairs long

• Different mutation rate than nuclear DNA No recombination but faster

mutation!• Scan loop for SNPs

Wikipedia File:Mitochondrial_DNA_en.svg



Science Background - SNPs

• Single Nucleotide Polymorphism– ACCTTGCGCGCTATA– ACCTTGGGCGCTATA

• Occur throughout the genome– Nuclear and mitochondrial DNA

• Are used as genetic markers

Wikipedia - Dna-SNP.svg‎



• Understanding Haplogroups



How to create a haplogroup

• 1. Identify multiple SNPs from multiple people• 2. Create Haplotypes• 3. Tag SNPs are identified – make identifying process easier

Person 1

Person 2

Person 3

Person 4



Interdisciplinary Learning Opportunities

• Migration of ancient peoples• Identification of students with their “deep ancestry”• Challenge ideas of race/ ethnicity• Immigration/ assimilation• Make science relevant• Link to social issues – current and historical

Very engaging!!

These are some of the 30 Tufts students who have worked on the HHMI project



Classroom Implementation – Medford High SchoolDay 1: Saliva collection Day 2: PCR Day 3: Gel electrophoresis

400 bp band

Send to Dolan – they sequence

Class results go on server

Students can access results

• Articulate students’ ideas about race– Race: The Power of an Illusion

• Compare mtDNA sequence to world population– GeneticOrigins.org– Bioservers



Ligation – Developed by Tufts UndergraduatesSaliva collection DNA purification PCR (mtDNA)

Gel electrophoresisLigationBiosensor readout

“T” “G”

Ligation No ligation

Control line



Ligation-based Assay

Mitochondrial DNA

sample #1

T

SNP specific probe

A

AAAA…AAA

G Mitochondrial DNA

sample #2

ASNP specific probe

AAAA…AAA

Common probe

Ligation!

No ligation

Common probe

B

B



Biosensor Readout

BTTT….TTTAAA.…AAAStreptavidin

• Students create the biosensors

• Control line – poly A oligonucleotides

• Experimental line – streptavidin

• If 2 probes ligate, 2 red lines appear –biotin binds to streptavidin *and* poly T beads bind to poly A tails

Control

Sam

ple

flow



Examples of Working SNPs on Biosensors

F-G10310A T-T10463C M-C10400T

G A G A T C T C C T C T83 54 48 15 21 2

AlleleSample ID

Haplogroup F? expect G at SNP site

Haplogroup T? expect T at SNP site

83 = no 54 = yes 48 = no 15 = yes 21 = no 2 = yes

Haplogroup M? expect C at SNP site



• Understanding Haplogroups



Running the Assay in the “Classroom”

• 7 high school juniors and seniors (Feb vacation – 2010)• 7 Post-Docs, graduate and undergraduate students• From start to finish: 9am-6pm• Students were eager to learn• Students had good ideas for simplification• Streamlining protocols – ready in late spring 2010

M-C10400T PrC

158 159 160 161 162 163 164 165

M-C10400T PrT

158 159 160 161 162 163 164 165



CO-OP Dissemination Plan

Developed by HS teachers & Tufts Staff

• Summer Workshop at Tufts • Equipment Lending Library

– Tufts maintains equipment• Scheduled Times for Experiments

– Google calendar• Tufts staff & undergrads help in classroom• Teacher feedback about experiments• Teachers networking between high schools• Generate new ideas – next year’s workshop

Resource Sharing Model



CO-OP

Medford High School

Malden High School

Josiah Quincy HS (Chinatown)

Somerville High School

Tufts University



Acknowledgements

• Funded by Howard Hughes Medical Institute Professor’s Award • Professor David Walt• Tufts Undergraduate student developers (see below)• Graduate student mentors (see below)• Post Doctoral mentors• Medford High, Malden High, Somerville High, Chinatown High



Interested in this project? • Sign up on our email list• Take a one page flyer about the project• Links to developed activities• We’ll be in touch!

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