HeadSTARTa Electrophoresis Kit J682

Directions for Use

Crime Scene:

Evidence to-date

The Victim: Margot Ellen Bota, a.k.a. "Ms. B", was found dead by her postman at the bottom of the staircase in her home. The residence showed signs of a struggle; however, nothing appeared to be missing - The victim was still wearing a diamond cluster ring. The back door was found slightly ajar with no signs of forced entry, indicating that Ms. Bota may have known her assailant. The coroner has ruled the cause of death as a broken neck, possibly caused by a fall. The body also exhibited multiple contusions and some abrasions, both caused by a physical struggle. Estimated time of death was 2:30 a.m. Background information indicates Ms. Bota to have had numerous illegal dealings.

Questioning revealed 2 prime suspects who were unable to substantiate their alibis for the approximate time of death.

Suspect 1: Richard Thomas Allan III. Council member who will be announcing his candidacy for Mayor next month. Last seen with the victim two years ago prior to pursuing a political career. Mr. Allan said he was at home sleeping at the time of death, but a witness, a transient, recognized Mr. Allan's photo and placed him near the vicinity of the crime scene around the time of death.

Suspect 2: Donald James Rody. Recently married the daughter of a wealthy, influential family; resides out of state. Well known "party boy". Mr. Rody inherited a large sum of money but went broke after only a few years. Mr. Rody's past is not known to his wife or her family. Mr. Rody was in town on business the night of the crime but claims to have been in his hotel room sleeping. Ms. Bota was Mr. Rody's companion prior to his marriage.

Evidence: Some brunette hairs were found entwined in the victim's ring. Blood was found under the victim's fingernails. Samples of blood were taken from both suspects and the victim. All evidence has been sent to the lab for DNA isolation and analysis.

 

HeadSTART Electrophoresis Kit

Technicians have prepared the DNA obtained from the crime scene for analysis by first isolating the DNA from the blood samples and then performing restriction digestions. Restriction digestions utilize enzymes that "cut" the DNA into many different sized fragments. These fragments can then be separated by gel electrophoresis and their restriction patterns analyzed.

 Like fingerprints, every individual's DNA is unique and can, therefore, be used for identification. By cutting several different samples of DNA with one enzyme, a wide range of restriction digestion patterns can be produced. Each individual sample will have its own distinct banding pattern and can easily be distinguished from all other samples. Your task is to take the digested DNA samples from the crime scene and perform restriction digestion analysis by electrophoresis (DNA fingerprinting). Note: The guilt or innocence of a person is at stake here, so take special care when handling the DNA and recording the lane position of each individual sample.

  • Prepare 10X TBE by dissolving the contents of the 10 X TBE Ready Pack in 500 ml of deionized water. Bring solution to a final volume of 1L with deionized water. Label as "10X TBE". Prepare 1L of 1X TBE by diluting 100 ml 10X TBE in 900 ml deionized water.

  • Weigh into a 250 ml beaker or flask 1 g of agarose (Code 0710). Add 1X TBE to a final volume of 100 ml.

  • Heat agarose solution just to boiling in a microwave oven or on a hot plate. Swirl solution and continue to gently heat until dissolved.

  • Pour gel in casting tray according to manufacturer's recommendations and insert comb. Allow gel to solidify for 20 to 30 minutes.

  • When completely solidified, place gel in apparatus and cover gel in 1X TBE and carefully remove comb. Once solidified, gels can be stored overnight in 1X TBE at 4°C. The buffer volume should be sufficient to completely cover the surface of the gel.

  • Pipette 10 l of each DNA sample per lane. Change pipette tips between loadings to eliminate chance of carry-over contamination.

  • HINT: Placing a strip of black paper under the wells will make the wells easier to see for proper loading.

  • Pay close attention to the dye as this will allow the sample to be visually tracked as it is loaded. For a perfect load, the dye should only be visible in the well. We recommend using the practice dye for a few loadings to familiarize students with proper loading technique. Care should be taken not to damage the bottom of the wells.

  • Cover the electrophoresis apparatus and apply voltage at about 5V/cm gel length (e.g. a 10 cm gel would be run at 50 V). Allow migration to continue until the red tracking dye is about 2 centimeters from the end of the gel. Turn the power off and remove the leads.

  • Carefully remove the gel and place in a 1X solution of DNA Stain (a 1X solution can be made by diluting 1 ml of the 100X DNA Stain in 99 ml deionized water). Allow to stain for 30 min. At this point the gel will be stained intensely blue with no DNA bands visible.

  • Carefully remove the gel and place in a 1X solution of DNA Stain (a 1X solution can be made by diluting 1 ml of the 100X DNA Stain in 99 ml deionized water). Allow to stain for 30 min. At this point the gel will be stained intensely blue with no DNA bands visible.

  • Destain the gel overnight in 200 ml deionized water. For best results, change the deionized water after about 1 - 2 hr.

  • Remove the gel from the destain bath and view either over a white light source or a white piece of paper (if a light source is not available).

 

Restriction digestion patterns (restriction fragment length polymorphism's or RFLP's) should be clearly visible in the destained gel. By comparing the RFLP's to one another, similar banding patterns should be visible. Match the banding patterns found in the evidence with the banding pattern found in the other samples. Analyze carefully the "Victim" DNA to ensure that the "Evidence" DNA did not come from the victim. Based on the RFLP results, determine which (if any) suspect is responsible for the crime.

 

211 Bis Avenue Kennedy

BP 1140

03103 Montluçon cedex - France

tel (33) (0) 4 70 03 88 55 

fax(33) (0) 4 70 03 82 60

interbiotech@interchim.com

 http://www.interchim.com

Vos contacts:

Etienne Boireau - Olivier Cadas - Olivier Alméras

hot line  33 (0) 4 70 03 73 06