Bio Lab Practical Exam

measuring absorbance: 1

labquest app, sensors, calibrate

measuring absorbance: 2

90 sec warm up, blank cuvette in, finish calibration

measuring absorbance: 3

sensors, data collection, time based mode

measuring absorbance: 4

red box > select wavelength, change absorbance

measuring absorbance: 5

cuvette in blank, play, 10 seconds > stop

measuring absorbance: 6

analyze, statistics, record avg (mean) absorbance

measuring absorbance: 7

cuvette 1 in cuvette 0 place, play, discard data

calculating pH: 1

probe into channel 1, rinse w/ distilled water, wipe w kim wipe

calculating pH: 2

submerge probe, wait 15 sec before data collection, play button > collect data for 10 sec

calculating pH: 3

stop button, analyze, statistics, record avg pH

calculating temperature: 1

temp probe into channel 1, stick probe/buffer into diff environments

calculating temperature: 2

play, collect data 10 sec, stop, analyze, statics, record avg temp

total solids

if solids mass at least 0.025 > after subtracting final - initial mass, proceed

BOD: 1

DO probe into channel 1, rinse tip w/ distilled water, wipe w kim wipe

BOD: 2

switch on probe to mg/L, submerge for 40 sec, collect data for 10 sec, stop

BOD: 3

analyze, statistics, avg DO concentration, record under initial dissolved oxygen

BOD: 4

still submerged, flip switch to %, press play, measure 10 sec, rec avg % saturation

Nitrates: 1

clamp probe, insert nitrate ISE to clamp vertically, uncap nitrate high standard, place o support stand directly under nitrate ISE

Nitrates: 2

lower probe clamp to submerge nitrate ISE in nitrate high standard (NOT touching bottom), soak for 30 min

Nitrates: 3

ISE sensor into channel 1 of vernier interface, sensors, calibrate, select nitrate ISE

Nitrates: 4

“calibrate now”, wait for live voltage stabilization, value 1 = 100, keep, swap nitrate high standard for rinsate beaker

Nitrates: 5

rinse, swap rinsate beaker for nitrate low standard, lower clamp so nitrate ISE submerged into low standard

Nitrates: 6

wait for live voltage stabilization, know value 2 = 1, keep, finish calibration, rinse ISE

Nitrates: 7

ISE tip into Nalgene bottle, probe submerged 60 sec, collect data 10 sec, stop collection, analyze, statistics, rec avg nitrate concentration

total phosphate: 1

calibrate spectrovis, connecct to labquest via USB, sensors, calibrate, spectrometer, 90 sec warmup

total phosphate: 2

blank in cuvette holder of spectrovis, finish calibration, ok, red box, change wavelength, enter given absorbance, ok

proteins

dependent on pH, temp, salt concentration of reaction mixture

buffers

aqueous solution mix, function to resist changes in hydrogenn ion concentration

stock solutions

concentrated solutions that last over a long period

dilution factor

concentration of dilute solution reduced compared to concentration of stock solution

serial dilution

stepwise dilution where stock solution for each dilution is dilute solution from previous dilution

spectrophotometer

machine measures absorbance or transmittance of pigmented solutions

standards (solution)

have known amount of material assayed, used to calculate amount in unknowns

standard curve

determine concentration of unknown

independent variable

x axis, not affected by other variables

dependent variable

y axis, influenced/changes in response to independent variable

discovery science

uses data or surveys of natural systems to discover patterns/correlations

hypothesis-driven science

uses scientific method to develop knowledge

descriptive statics

describe/summarize data

-mean, median, mode measure central tendency

-range/variance/standard deviation measure dispersion

inferential statistics

make estimates/draw conclusions/inferences about population based on sample (T-Test)

temp effects on WQI

-cold water = more dissolved gas/DO

-temp changes indicate source of thermal pollution

-increased temp increases photosynthesis

-organisms stressed, lowering resistance

thermal pollution

caused by human activities

runoff

increases overall temp of water

shade

important to stream health, blocks too much direct sunlight

pH effects of WQI

algal blooms, industrial processes, sulfide sediment oxidation, rainfall

total solids effects on WQI

-soil erosion

-runoff

-plankton/animal and plant matter

-dissolved solids dehydrate organisms

-high total solids > cloudy water > less photosynthesis

factors affeting DO concentrations

temp, stream flow, air pressure, aquatic plants, decaying organic matter, human activities

BOD effects on WQI

-Oxygen replenished faster than used

-too much organic material to decompose = high BOD

-initial DO - final DO = BOD level

Nitrate effects on WQI

-increased by fertilizers, decomposition, waste, runoff, etc.

-high levels cause eutrophication

Total phosphate effects on WQI

-high levels associated w decreased DO/increased BOD

-added by humans

Fecal coliform bacteria, WQI

-indicated other pathogens from feces

-caused by leaking sewer systems, polluted runoff, etc.

-can survive wo oxygen

Turbidity, WQI

-increase w stream flow, soil erosion, runoff, etc.

-caused by bottom dwelling organisms, organisms, plankton

Conserved DNA region

-nucleotide seq w little/no variation across species, remains constant throughout evolution

Bacteria DNA barcoding

-agar as growth medium

-no buffer, prokaryotic

DNA purification/isolation: 1

Heat sample to 94 degrees celcius for 5 min, purify DNA, centrifuge Lysate

DNA purification/isolation: 2

DNA binds column, wash buffer clears everything else away (4x), nuclease free water added to column/centrifuged, DNA pulled through column/isolates

PCR

-Denaturing: DNA in microcentrifuge for 1 min at 95 degrees celcius

-annealing

Master mix

-DNA polymerase, dNTPs, buffer, forward/reverse primers, water

-reduced pipetting errors/time running reaction

Gel electrophoresis: 1

-separate, identify, purify DNA frags

-DNA migrates to wards + charged electrode, smaller move faster through matrix

-Agarose/buffer cool/hardening jello substance

gel electrophoresis: 2

-blue gel dye added, EtBr added for frag visualization

-gel run at 175 volts, turn off when migrated 5 cm

gel analysis: presence of ladder

gel made properly, electrophoresed in correct direction, stained by etbr

gel analysis: no bands including ladder

etbr not added

gel analysis: ladder visible but bands not

part of taq polymerase degraded maybe

gel analysis: no sample band

one or both primers not added to master mix, DNA extraction recovered little/no genomic DNA

gel analysis: no band in bacteria

did not properly inoculate bacteria culture

gel analysis: multiple PCR bands

non-specific primer binding, sample contamination, primer added to multiple sample. master mix