Objectives: 09/04/2014 Lab Meeting Article Info Finish lab 3a Discuss lab 3b Lab notebooks will be collected at the end of class tomorrow

Bellwork: 11/01/ mL = _________ L _______________ 2. P-10/20/100/1000 measure what volumes?

Today: 11/04/ How would you calculate the mass required to make the following solutions: 5 mL of 2.5% glucose solution: 2000 mg = ___________g 5L = ___________mL 6.3cm = _________mm

Using the Spec: 1.Turn the instrument on and allow it to warm up for 15 minutes. 2.Set your spec to the desired wavelength. 3.Use a kim wipe to clean off any finger prints, oil, debris, etc., from the outside of your test tube. 4.Insert the standard test tube (deionized water) and blank the spec. 5.Insert your test sample and read/record both %transmittance and absorbance. 6.Repeat steps 4 and 5 for each testable sample.

A) Metric System IS: International system of measurement PrefixSymbolFactor kilok1000 hectoh100 (base unit) no prefix----1 decid0.1 centic0.01 millim0.001 micro µ (10 -6 ) nano n (10 -9 )

A) Metric System

To easily convert between prefixes we can use the BS rule

B) Measuring Volumes The main units we measure volumes in are microliters, milliliters and liters 1000 µL = 1 mL = L  *know these conversions!!!!!  Practice: 1) 125 mL = _________ µL 2) 33 mL = _________ L 3) 4 L = _________ mL 4) 200 µL = _________ mL 125, ,

C) How do we measure volume in the lab? Graduated cylinder: Measure more than 10 mL Pipet: measure between 0.5 mL &10 mL Micropipet: less than 1 mL

C) How do we measure volume in the lab? Given the following volumes, what instrument would you use to measure that volume: 5) 100 µL 6) 25 µL 7) mL 8) 56 mL 1) 100 mL 2) 25 mL 3) 5 mL 4) 10 mL Graduated Cylinder Graduated Cylinder Pipet Pipet or Graduated Cylinder Micropipet

D) How to use a pipet Draw this: In order to use a pipet you much also have a pipet pump: Blue: 1- and 2-mL pipets Green: 5- and 10-mL pipets Red: 25- and 50-mL pipets

Roll the gear so the top raises, this will draw up the liquid into the pipet Roll the gear the opposite direction to expel the liquid DO NOT push the top of the pipet pump – this may break the pump When selecting a pipet to use remember to use the smallest pipet as possible to be as precise as possible

E) How to use a micropipet Used to measure amounts less than 1 mL 3 sizes of pipets: P100.5 µL to 10 µL P µL to 100 µL P µL to 1000 µL Each micropipet has only a certain amount of precision Make sure you pay attention to this!

E) How to use a micropipet These are the readings for micropipets Main difference between 20 and 10 is the top limit, it still has the same unit places on the display

E) How to use a micropipet P10/P20

E) How to use a micropipet P100/P200

E) How to use a micropipet P1000/P2000

F) Making Solutions One of the most essential skills Labs often buy the solid form of a compound because it is often cheaper and many times multiple concentrations are used. Almost all reactions involving proteins and nucleic acids occur in an aqueous solutions. Solution contains 2 parts: Solute – what is being dissolved Solvent – what us doing the dissolving

F) Making Solutions To prepare a solution: 1. Measure out solid mass of solute needed 2. Add solid to container with a little water 3. Dissolve solid 4. Add enough water to reach desired volume

Bellwork: 09/23/2014 You need 50 L of D-Fructose for a bioreactor. This D-Fructose must be at a concentration of 12 mg/mL. How many kilograms of D-Fructose do you need?

G) Concentrations A measure of how many particles are in a certain volume Normality made be used for acids & bases only Common Units of Concentration ConcentrationCommon Units of Measurement mass/volumeg/L, mg/mL, µg/mL, µg/µL % mass% MolarityM (mol/L), mM (mmol/L), µM (µmol/L)

H) Preparing Mass/Volume Concentration Mass/Volume Concentration Equation Conc. desired x volume desired = mass of solute (g/mL)(mL)(g) Example: How do you prepare 100 mL of 0.05 g/mL solution of hemoglobin? 1)Use the equation to determine mass of solute g/mL x 100 mL = 5 g hemoglobin 2)Give directions for making solution: Measure out 5 g of hemoglobin and pace into a 100 mL flask, add about mL of dH 2 O. Dissolve all hemoglobin then add enough water to make 100 mL of solution.

H) Preparing Mass/Volume Concentration Mass/Volume Concentration Equation Conc. desired x volume desired = mass of solute (g/mL)(mL)(g) Practice: How do you prepare 150 mL of 2 mg/mL gelatin? 1)Use the equation to determine mass of solute. 2 mg/mL x 150 mL = 300 mg gelatin = 0.3 g gelatin 2)Give directions for making solution: Measure out 0.3 g of gelatin and dissolve in a little bit of water. Add enough water to make 150 mL of solution.

% Mass/Volume Concentration Equation Convert % to decimal Decimal % x volume desired = mass of solute (g/mL)(mL)(g) I) Preparing % Mass/Volume Concentration Example: How do you prepare 50 mL of 10% NaCl solution? 1)Use the equation to determine mass of solute g/mL x 50 mL = 5 g NaCl 2)Give directions for making solution: Measure out 5 g of NaCl and dissolve in a small amount of water. Add enough water to make 50 mL of solution. 10% → 0.10 g/mL

I) Preparing % Mass/Volume Concentration % Mass/Volume Concentration Equation Convert % to decimal Decimal % x volume desired = mass of solute (g/mL)(mL)(g) Example: How do you prepare 60 mL of 10% glucose? 1)Use the equation to determine mass of solute g/mL x 60 mL = 6 g glucose 2)Give directions for making solution: Measure out 6 g of glucose and dissolve in a small amount of water. Add enough water to make 60 mL of solution. 10% → 0.10 g/mL

09/17/2014 Your background must be complete prior to starting the lab. Everyone must wear gloves, lab coat/apron, and eye protection We are using Methylene Blue instead of Pyronin Y

09/17/2014 – Lab Changes We are using Methylene Blue instead of Pyronin Y You will microwave your agarose to melt it. You will use TAE buffer instead of TBE buffer Microfuge Tube Symbols: Bromophenol Blue – “B” Orange G – “G” Methylene Blue – “M” Safranin O – “O” Xylene Cyanol “X” Unknown – “?”

General Critiques You need to know what you are doing before you do it. Read your instructions and ask questions prior to starting. Do your math first! Your peers are a very good resource, don’t follow me around asking for help when you have 20 peers that can answer your question. Don’t use the analytical balances unless you need to! Remove scoops and close containers once your are done using them

Power Supply Settings 100 V 50 minutes 35 amps Make sure the wells are covered in TAE buffer Do not start the power supply until four boxes are connected

Bellwork: 09/19/2014 Get started! If you ran your gel yesterday, work on entering your data into your lab notebook and writing your conclusion. If you did not run your gel yesterday, hustle to get the gel loaded and start the power supply. Once all the gels are running we will start our mini presentations

Bellwork: 09/22/2014 How would you prepare 10 mL of 50 mg/mL CuSO 4 5H 2 O Formula: Total Volume x Concentration = mass of the solute

Lab 3e: Making Solutions of Differing Mass/Volume Concentrations The regular lab report format is not needed for this lab, but the following should be documented in your lab notebook: Tables 3.8, 3.9, and 3.10 A print out of step “h” on page 47 Questions 6, 10, and 14 on page 49 Be sure to take two sets of spec readings! PPE – gloves, goggles, lab coat/apron

Objectives: 09/26/2014 Complete through lab 3e and turn in your lab notebook for grading Science article presentations Begin work on Lab 3f

Bellwork: 09/29/ If you were pouring a gel for DNA agarose gel electrophoresis, where would you place the comb? Sketch a picture and explain your reasoning. 2. Make sure the data you are recording is actually your data, not an ideal from a recipe, and be concise in your writing

Lab 3f Requirements The regular lab report format is not needed for this lab, but the following should be documented in your lab notebook: Tables 3.11, 3.12 Show your math for one tube in table 3.11 Run Part IV twice – Once with the professionally made Biurets and once with yours Do not do the spec/absorbance steps PPE – gloves, goggles, lab coat/apron We will not be doing labs 3g/h, but you are responsible for knowing the math within those labs

Quick & Easy Bellwork: 1. How does gel electrophoresis work? 2. Give an example of when a spectrophotometer would be used, but the solution used to blank the device would not be water

J) Preparing solutions using Molarity Molarity Units: mol/L mole = the number of particles in a substance that is equal to the mass (in grams) on the periodic table Ex: 1 mole NaCl = _____ g 1 mol of any substance will not be the same as a different substance! 1 mole Na = 23.0 g1 mole Cl = 35.5 g 1 mole NaCl = 23.0 g g = 58.5 g 58.5

J) Preparing solutions using Molarity Practice: Solve the following. 1) 1 mol C 6 H 12 O 6 = ________ g 2) 1 mol CaCl 2 = ________ g 180.0g and 111.1g are called the molecular weight (MW) or formula weight of its substance (12.0 x 6) + (1.0 x 12) + (16.0 x 6) (40.1 x 1) + (35.5 x 2)

J) Preparing solutions using Molarity Molarity Concentration Equation Volume wanted x molarity x MW = mass of solute (L)(mol/L)(g/mol)(g) Example: How many grams are needed to make a 75 mL solution of 2M CaCl 2 ? L x 2M x g/mol = 16.67g CaCl 2 Practice: How do you prepare 20 mL solution of 0.25 M NaCl? L x 0.25M x 58.5 g/mol = g NaCl Measure out 0.293g of NaCl and dissolve in enough water to make 20 mL of solution.

What mass of calcium chloride (CaCl 2 ) is needed to make 125 mL of 0.55 M CaCl 2 solution?

1 st convert 125 mL into L (molarity = moles/liter) 125 mL = L Then use the molarity concentration equation

Volume wanted x molarity x MW = mass of solute L x 0.55 M CaCl 2 x 111 grams/mole = 7.63 grams of CaCl 2 and add solvent up to 125 mL

K) Dilutions Buying concentrated solutions can save money If you need a very dilute solution starting from a more concentrated solutions may be required

K) Dilutions Dilution Equation C 1 V 1 =C 2 V 2 C = ConcentrationV = Volume 1 & 2 stand for different solutions Concentrations can be in any unit as long as both are the same, same with volume Example: How would you prepare 1 L of 1mg/mL protein solution from 100mg/mL concentrated stock? C 1 = 1 mg/mLV 1 =1LC 2 =100 mg/mLV 2 =? (1 mg/mL)(1L) = (100 mg/mL)(V 2 )V 2 =10mL Measure out 10 mL of protein stock solution and mix in enough water to make 1 L of solution.

K) Dilutions Dilution Equation C 1 V 1 =C 2 V 2 C = ConcentrationV = Volume 1 & 2 stand for different solutions Concentrations can be in any unit as long as both are the same, same with volume Practice: How do you make 200 mL of 10mM CaCl 2 from 75mM CaCl 2 solution? C 1 = 10 mMV 1 = 200 mLC 2 =75 mMV 2 =? (10 mM)(200mL) = (75 mM)(V 2 )V 2 =26.6mL Measure out 26.6 mL of CaCl 2 and mix in enough water to make 200 mL of solution.

Get to Work! Changes to the Lab: Step 3 (page 56) – You do NOT need to graph your data. The trend(s) should be obvious through a data chart. For your conclusion: Possible errors Results with evidence Practical use for this technique Thinking like a Biotechnician, questions 1, 2, 3 A summary of page 58 – why is pH biologically important? Etc.

Get to Work! For your background section: Explain: mole, molarity, Avogadro's number, and molecular weight Write out the molarity concentration equation (include all units) Solve the following, showing all of your work: What is the molarity of a solution containing grams of RuCl 3 in enough water to make 1.00 L of solution? How many grams of KMnO 4 are required to prepare 1.0 L of a solution of 1.5 M KMnO 4 ?

Get to Work! For your background section: Explain: mole, molarity, Avogadro's number, and molecular weight Write out the molarity concentration equation (include all units) Solve the following, showing all of your work: What is the molarity of a solution containing grams of RuCl 3 in enough water to make 1.00 L of solution? How many grams of KMnO 4 are required to prepare 1.0 L of a solution of 1.5 M KMnO 4 ?

Unit Equations: Mass/Volume Concentration Equation Conc. desired x volume desired = mass of solute (g/mL)(mL)(g) % Mass/Volume Concentration Equation Convert % to decimal Decimal % x volume desired = mass of solute (g/mL)(mL)(g)

Unit Equations: Molarity Concentration Equation Volume wanted x molarity x MW = mass of solute (L)(mol/L)(g/mol)(g) Dilution Equation C 1 V 1 =C 2 V 2 C = ConcentrationV = Volume 1 & 2 stand for different solutions Concentrations can be in any unit as long as both are the same, same with volume