Fall 2021 BIOL 115 Lab Manual - Google Docs - Copy PDF

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BIOL 115 LAB MANUAL Dr. Miranda A. Kearney          1

TABLE OF CONTENTS ** This table of contents contains links to each section listed below. If you click on the section you are interested in viewing, a link will come up. Click this link and the doc will automatically navigate to the proper page.** Fall 2021 Tentative Schedule and Assignment Due Dates

4

Research Project Overview

6

Methodological Proposal Proposal submission link The Experiment Collecting Weekly Descriptive Data & End of Experiment Data Descriptive data collection protocols

9 9 9 14 14

Shoot size Number of leaves

14 14

Average leaf size End of Experiment data collection protocols

14 14

Root Scans Measuring Fresh Weight Biomass and Root:Shoot ratio

14 15

Archiving Soil Samples

15

Practice Analysis- ImageJ for Measuring Roots Root importance submission link

16 16

Practice Analysis- Graphing Data Plant data practice graphs/figures submission link

17 18

Practice Analysis- Statistical Analyses Practice statistics submission link

18 22

Assignments

22

Homework Assignment 1- Time Management & Planning Homework Assignment 2- Methods Section

23 23

Reflection Essay Instructions

24

Appendix 1: A Guide to Data Management Using MS Excel

24

Appendix 2: A Guide to Simple Statistics

35

Appendix 3: Rubrics and Guides Presentation Guide

40 40

Presentation Rubric

40 2

Elevator Speech Guide

41

Elevator speech Rubric Scientific Poster Guide

42 43

Scientific Poster Rubric Research Paper Draft Rubric

43 44

Final Research Paper and Discussion Section Rubric

44

Appendix 4: Contribution Index

44

Appendix 5: Team Contract

44

Appendix 6: CSE Citation Format Guides and Resources

44

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Fall 2021 Tentative Schedule and Assignment Due Dates Lectures are on Fridays from 8:30-9:30am in LH1. It is expected that you will be at lecture each week. We will cover the following topics in lecture across the semester and each of these topics will cover content that will be directly important/relevant to the research being done in lab: 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12.

Course Overview, Syllabus Review, Process of Science Descriptive Statistics Types of Data Graphing- Choosing the right graphs for the right data Making Sense of Scientific Papers (MSSP): Method Sections MSSP: Results Sections (including figures and figure legends) ANOVAs and MANOVAs Elevator Speeches + Workshop MSSP: Scientific Literature MSSP: Discussion Sections MSSP: Titles & Abstracts Wrap Up- Research and where do you go from here? Week of:

1

Lab T, W, Th in S2 153C, 154, 155 NO LABS

Aug 23 (classes start on T 8/24)

2

30

3

Sep 6 (no classes 6-8)

Read Lab Manual “Research Project Overview” including the 2 articles (links in The Project), “Introduction” and “The Project”. Stop at “The Tools” Team Placement & Team Contract Planning: Software Introduction & Downloads Project Design Begin Research Outline/ Proposal NO LABS TUESDAY & WEDNESDAY THURSDAY LABS MEET! Continue Drafting Research Outline/ Proposal Equipment Introduction & Leachate production

4

Pre Assessment Due by 5pm Friday NO THURSDAY LABS

13 (no class Th 9/16)

Continue Drafting Research Outline/ Proposal Equipment Introduction & Leachate production

5

20

W0

6

27

W1

7

Oct 4

W2

Proposal Due by 5pm Friday for feedback Begin Research Project (RP)- Prepare rhizotrons Practice Analysis- ImageJ for Measuring Roots Homework 1 Due Continue RP- Collect Descriptive Data, Apply Treatments Practice Analysis: Graphing Data Presentation 1 Due CATME 1 Due Continue RP- Collect Descriptive Data, Apply Treatments Presentation 2 Due Methods Section (Homework 2) Due

4

8

11 (No classes 14-18 & 8/12 is a Th)

W3

9

18

W4

10

25

W5

11

Nov 1

12

8

NO LABS & NO LECTURE TAs and Dr. K WILL ADD TREATMENTS THIS WEEK Continue RP- Collect Descriptive Data, Apply Treatments Practice Analysis: Statistical Analyses Research Project Final Data Collection Practice Analysis: Statistical Analyses ctd. CATME 2 Due Begin Data Analysis & Poster Building Elevator Speeches Due Continue Data Analysis & Poster Building Research Paper Draft Due

13

15 Continue Data Analysis & Poster Building

14

22 (No classes 24-28)

15

29

NO LABS & NO LECTURE Poster Presentations

16

Dec 6 (Last day of classes 12/10)

Final Poster & Poster Talk Due Future Research Discussion and Presentation (no outside prep needed, done fully during lab time)

17

13 (Finals Week)

Reflection Essays Due by 5pm Friday Final Research Paper & Individual Discussions & Assessments (LCAS & Post) All due by 5pm Monday 12/13

5

Research Project Overview INTRODUCTION Welcome to BIOL115 and the Kearney Team! This semester you will be working on a novel research project. That means that we (I and the TAs) do not know what you will find. No one else has done this research before and people outside of this classroom (other scientists, stakeholders, and the general public) would be reasonably interested in what our results show. We are excited to work with you on this project! We are working within a tight time-frame to complete a full research project (one semester), so Dr. Kearney has already done some of the work for you. She has identified the novel questions we are going to find answers to this semester. You may be really interested in the project or it may not exactly be centered within your favorite area of biology. Either way, that’s OK. (But Dr. K will use some of our lecture time to try and pique your interests in this area of study!) This course is an opportunity for you to immerse yourself in the scientific process. You will learn and practice skills that will help you as you progress through University and beyond (for specifics- check out the learning goals and outcomes section in the course syllabus). THE PROJECT Research begins with a question and often multiple questions are addressed in one study. The research we do this semester will explore the following questions: 1. Does leachate from disposable face masks (DFMs) impact survival in plants? 2. Does leachate from DFMs impact growth and development of aboveground plant structures (shoots) or belowground plant structures (roots)? 3. Does a contaminant mixture (leachate from DFMs and a common pharmaceutical contaminant (ibuprofen)) impact survival in plants?

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4. Does a contaminant mixture (leachate from DFMs and a common pharmaceutical contaminant (ibuprofen))impact growth and development of aboveground plant structures (shoots) or belowground plant structures (roots)? We will investigate these questions by germinating seeds, treating the seedlings weekly for 5 weeks with experimental solutions (including a DFM leachate solution made following a modified protocol from (please read it!): Sullivan GL, Delgado-Gallardo J, Watson TM, Sarp S. 2021. An investigation into the leaching of micro and nano particles and chemical pollutants from disposable face masks - linked to the COVID-19 pandemic. Water Research. 196 (117033). https://doi.org/10.1016/j.watres.2021.117033 Then collecting and analyzing data on mortality and growth and development of plant above and belowground structures. We will be able to examine root structure by using a device called a rhizotron (see image above in the introduction) and using a slightly modified approach based heavily on this protocol (please read it!): Cassidy ST, Burr AA, Reeb RA, Melero Pardo AL, Woods KD, Wood CW. 2020. Using clear plastic CD cases as low-cost mini-rhizotrons to phenotype root traits. Applications in Plant Sciences. 8(4): 1–7. https://doi.org/10.1002/aps3.11340 Before we start the experiment, during the lab portion of the course we will take time to familiarize ourselves with the protocols, equipment, and tools that everyone will need to be able to complete this research. We will also form research teams that will tackle the project together. Science is rarely done alone and this class is no exception. As a research team, the first thing you will tackle is writing a methodological proposal (described in more detail later in this section) that describes exactly how your research team will execute this research project. THE TOOLS Lab time has been set aside for you to acquire each of these programs. Please do not try to access them before your TA goes over this with you. We will use the following computer programs to complete the research for this course: Program

Details

GSuite

Binghamton University is a google campus. You all have access to google drive and the suite of google programs through your binghamton.edu address. As a team, you should create a shared drive or a 7

shared folder in which you will collaborate as a group. The first document that should go in this drive is your completed Team Contract. Save a copy to your shared drive and fill it out thoroughly! MS Excel

MS Office 365 is available to all students for free through Binghamton University. Visit: https://www.binghamton.edu/its/about/organization /technology-support-services/hardware-software/o ffice-365.html and follow the instructions to download There is a guide to data management in excel in Appendix 1 of the lab manual. You can also find many useful tutorials by simply googling what it is you want or need to do.

Plot.ly

Use this link to sign up for a free account. There are tutorials available here or you can play around with the program yourself and/or with your teammates.

ImageJ (root analysis software)

Visit: https://imagej.nih.gov/ij/download.html and follow the instructions to download directly

GIMP (GNU Image Manipulation Program) (optional)*

Visit: https://www.gimp.org/downloads/ and follow the instructions to download directly

PAST (PAlaeontological STatistics)

Visit: https://www.nhm.uio.no/english/research/infrastruc ture/past/ and follow the instructions to download *Mac users: if you get a message “developer can't be verified” follow these instructions to open: https://support.apple.com/guide/mac-help/open-amac-app-from-an-unidentified-developer-mh40616 /mac

THE PROPOSAL You all should have read the first part of this Research Project Overview including the 2 articles (links in “The Project” section), “Introduction” and “The Project” sections. If you haven’t DO IT NOW, then as a team begin to hammer out your methodological proposal. The purpose of this proposal is to completely outline the experimental design for your research project including all of the specifics from beginning (set up) to end (analysis). Every team will have 12 minirhizotrons available to use plus soil and seeds. You will need to make your DFM leachate solutions but we will provide you with the pharmaceutical/ibuprofen solution. Keep in mind that about 10 ml of treatment solution is enough to moisten a minirhizotron without overdoing it. 8

Methodological Proposal Your methodological proposal should include the following sections: Rationale: Why is this project important to do? For this section, you need to write a paragraph based on what you know already and based on some preliminary background information you gather as a group. You will be expanding on this as we move forward and we don’t expect you to have a really solid grasp on this yet. Questions: List out the questions you intend to address with this project (see above). Hypotheses: These may be refined as you learn more background about our research project. Typically, a scientist wouldn’t begin an experiment until they had a solid understanding of what’s already known about the topic they are studying. However, we have a very short window of time to complete our research, so we will be doing some things a little out of the typical order. Materials and Methods: This section is used to describe what you intend to do, so that a person reading it would be able to set up and run (basically recreate) your experiment. To make this possible, you need to include all of the parts of the procedure. In addition to the actual set up and execution of the experiment, this also includes how your data will be analyzed (including any computer programs you will use), what statistical test(s) will be performed, etc.. You may also make and include figures or schematics that would help a reader fully understand your experimental set up. In short, it should be a full detailed description of what you plan to do. It should follow a logical order, use full sentences, paragraph form, and future tense and should NOT be written as a directive/instruction manual. It will be helpful to attend Dr. K’s Methods lecture and use your notes from it to help you with your proposal’s Methods section Proposal submission link Submit your proposal HERE by the due date provided in the course schedule. Please submit 1 doc (preferably a google doc) per team and include the names of all team members that contributed in that doc. This will not be graded, but you will receive feedback from your TA that you will be able to use to improve content for the graded methods section homework and the research paper draft.

The Experiment

9

THE RESEARCH OPTIONS Disposable face masks (henceforth DFMs) come in a variety of colors and patterns. For this research project, Dr. Kearney has chosen 6 different facemask styles to investigate. Each lab section has a max of 6 research teams and each team will be assigned (or choose) one of the styles to work with. Within a single lab section, each project will be unique, so there will be no repeats of DFM styles. Specifically we will make leachate from the following 6 DFM styles: Style

Image

Source

1. Traditional

https://www.amazo n.com/DisposableFace-Masks-Breath able-Comfortable/d p/B084TQKLCC/ref =sr_1_3?dchild=1& keywords=disposab le+face+masks+50 +pcs&qid=1628090 441&sr=8-3

2. Solid Black

https://www.amazo n.com/Black-Masks -Breathable-Stretch able-Elastic/dp/B08 DY96SSF/ref=sr_1 _4?dchild=1&keyw ords=disposable+fa ce+masks+50+pcs &qid=1628090441& sr=8-4

3. Leopard

https://www.amazo n.com/Leopard-Dis posable-Medical-Br eathable-Earloop/d p/B08LYTQFH6/ref =sr_1_19?dchild=1 &keywords=dispos able+face+masks+ printed+50+pcs&qi d=1628090304&sr= 8-19

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4. Camo

https://www.amazo n.com/YLY-Individu ally-Packaged-Disp osable-Camouflage /dp/B08QYV9RP6/r ef=sr_1_15?dchild= 1&keywords=dispo sable+face+mask+ pack+60&qid=1628 091783&sr=8-15

5. Holiday

https://www.amazo n.com/Christmas-Di spos%E1%BA%B5 ble-Bre%E1%BA% A1th%D0%B0ble-C %C3%B2r%C3%B 2n%C3%A2v%C3 %ADr%C3%BAs-Pr ot%C8%A9ct%E1 %BB%8Bo%C3%B 1/dp/B08QCXF979/ ref=sr_1_14?dchild =1&keywords=disp osable+face+mask s+christmas+print+ 50pcs&qid=162809 0665&sr=8-14

6. Rainbow

https://www.amazo n.com/DIOLV-Christ mas-Disposable-Br eathable-Holiday/d p/B08MXH2DYZ/ref =sr_1_25?dchild=1 &keywords=dispos able%2Bface%2Bm asks%2Bfestive%2 Bprint%2B50pcs&qi d=1628090927&sr= 8-25&th=1

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THE SETUP

Image Source: https://gardenerspath.com/plants/vegetables/grow-leaf-lettuce/

Lettuce seeds are SMALL! Setting up Rhizotrons Follow this link to a visual guide for how to prepare and seed your rhizotrons. This video from Spring 2021 may also be helpful. Use the above guides to set up 12 rhizotrons. Divide up tasks evenly among teammates. Important notes: 1. DO NOT ALLOW THE WATER SLEEVE OR THE DARKENING SLEEVE TO BLOCK THE OPENING OF THE RHIZOTRON 2. MAKE SURE THE SEED IS PLANTED NO MORE THAN 2CM BELOW THE OPENING AT THE TOP OF THE RHIZOTRON Labeling Rhizotrons Once you have set up all 12, you will need to label your rhizotrons. Each team has chosen the DFM style they will work with and will also work with the common pharmaceutical contaminant. There is also a control (water). You now need to label each of your rhizotrons with the DFM leachate they will be treated with for the duration of the experiment. Write this information directly on the darkening sleeve. Specifically, label the rhizotrons along the 2 “spines” AND on the front side. Stay consistent as to which spine you label for every rhizotron. See the “ORIENTATION” slide in the visual guide if you are unsure what is meant by “spine” and front. ● ●

4 rhizotrons- Control 4 rhizotrons- DFM # 12

●

4 rhizotrons- DFM # + pharmaceutical

Treating Rhizotrons For week 0 (W0- see tentative schedule), you will add 10 ml of water to each of your rhizotrons to allow the seed to germinate BEFORE we begin the experiment. The following week (W1) teams will begin their experiments. To treat your rhizotrons, you will use the correct syringes to add 10 ml of solution to the corresponding labeled rhizotron. Each team has 3 treatments and 12 rhizotrons so 4 replicates for the control, 4 for the DFM leachate, and 4 for the multi-contaminant (DFM + pharmaceutical) treatment. However, when we do any data analysis in the course, we will be using the FULL CLASS DATASET (since 4 replicates is not a strong enough sample size, this concept will be discussed extensively in lecture). ● 4 rhizotrons- Control (Add 10ml of control RO water to each rhizotron) ● 4 rhizotrons- DFM # (Add 10 ml of DFM leachate solution to the corresponding rhizotrons- make sure to gently mix the solution before transferring to the corresponding rhizotrons) ● 4 rhizotrons- DFM # + pharmaceutical (Add 5ml of mixed DFM leachate and 5 mls of provided pharmaceutical solution to the corresponding rhizotrons) Positioning Rhizotrons for Optimizing Growth The rhizotrons must be set up in light and angled at an approximate 45° angle so that the gravitropism response ensures that as the seed germinates and the plant starts to grow, the roots will grow up against one side of the rhizotron. The rhizotrons may be “stacked” on top of each other. Important Notes: 1. DO NOT COVER THE OPENING WHERE THE SEEDLING WILL EMERGE 2. MAKE SURE THE OPEN SPACE AT THE TOP OF THE RHIZOTRON FACES UPWARDS. See the “ORIENTATION” slide in the visual guide.

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Collecting Weekly Descriptive Data & End of Experiment Data Descriptive data collection protocols Descriptive data including mortality data will be collected weekly during the experiment and at the end of the experiment (after scanning rhizotrons but before harvesting all plant material for biomass measurements). This data will be useful in examining the progression of growth of your seedlings over the course of the experiment. The variables we will measure for each of the plants in your rhizotrons are: 1. Shoot size a. Use a ruler or calipers to measure the length of the longest leaf from the “ground” (top of the soil) to the tip of the leaf. 2. Number of leaves a. Count each leaf that is visible on the plant. 3. Average leaf size a. Use the ruler (or calipers) to measure the size of each leaf you counted. Be gentle so as not to break or injure the plant!...