INTRODUCTION TO NEOGEN DNA TESTING PROCEDURES

Transcription

1 INTRODUCTION TO NEOGEN DNA TESTING PROCEDURES 1. Order Test Sample Supplies from 2. Print and Fill out a DNA Request Form from under the FORMS tab (testing sample supplies must be in hand before submitting a DNA Request Form). 3. Return DNA Request Form, with the Barcode ID written clearly, and payment to the TLBAA Office. The Barcode ID must be correct and must match the Sample container to the animal being tested. 4. Once the DNA Request Form reaches the TLBAA Office, a Work Order Form will be generated by TLBAA. All test samples must be submitted with a Work Order Form. Any samples that arrive at Neogen without this work order form will not be identified. 5. When you receive your Work Order Form and order details. Ensure that the Barcode ID and Animal Information matches on your test sample and Work Order Form. DO NOT MAIL TEST SAMPLES TO TLBAA 6. Mail Test Samples, with the Work Order Form, to NEOGEN. 7. You will be notified when results are complete, and the TLBAA animal registry file will be annotated. TEST SAMPLES MUST BE MAILED TO NEOGEN. DO NOT SEND TEST SAMPLES TO TLBAA. ALL TESTING SAMPLE SUPPLIES MUST BE PURCHASED THROUH

2 HOW TO ORDER DNA TESTING SUPPLIES WITH IGENITY by NEOGEN Step 1: Go to Step 2: Select Order Sample Supplies from the menu on the left-hand side. Step 3: Select your preferred Sample Type and desired quantity. -Choose from: -Blood Cards...($0.50 per card/10 card minimum order) -Hair Cards..($1.00 per card/10 card minimum order) -Tissue Sampling Unit ($21.00 for 10 unit pack/ Applicator Gun $45.00 each) Step 4: Fill in Shipping Information. Step 5: Provide for Order Confirmation. Step 6: Review Your Order. Step 7: Submit. Step 8: Call (1-800-Igenity) to pay via credit card OR Mail payment to: Igenity 4131 N. 48 th St. Lincoln, NE DO NOT MAIL ANY TEST SAMPLES TO TLBAA. For all questions and concerns call TLBAA at (817) For all shipping concerns call Igentiy

3 Igenity Hair Sample Instructions 1. Pull (do not cut) hair from the tail switch, not tail head. Pull hair in the opposite direction from which the hair is laying. This results in less breakage than pulling straight out from the tail. 2. Make sure hair is free of fecal material and dirt. 3. Roots must be clearly visible. Approximately 30 hair roots are needed. For animals with finer hair, a minimum of 50 to 60 hairs is desired. For animals younger than three months, blood is recommended since hair roots will likely not be visible. 4. Open the collector and peel back the plastic cover. Insert the root end of hairs in the middle of the collector. Seal the plastic cover over the hair and then close the collector cover tab. 5. Trim excess hair extending from the collector. Write animal ID and other information in the spaces provided. 6. Properly record animal IDs on the collector cards and correctly enter that information on your testing order form to ensure you are matching the right DNA to the right animal. Neogen: GeneSeek Operations 4131 N. 48th Street, Lincoln, NE igenity.support@neogen.com Neogen Corporation, Neogen and Igenity are registered trademarks of Neogen Corporation, Lansing, Michigan. AG

4 Igenity Blood Sample Instructions Two to three drops of blood are needed on the collector portion of the card. This can be collected from anywhere on the animal. Two locations for easy-to-find blood vessels are the ear and the underside of the tail. 1. Appropriately restrain the animal. Locate a blood vessel, visually or by palpation. Clean the area so the sample is not contaminated with dirt or manure. Use a clean needle or lancet for every animal. 2. Blood can also be sampled with a syringe or blood tube from the vein on the underside of the tail. This may be easier than sampling blood from an ear vein that is covered with long, thick hair. 3. Collect two to three drops of blood on the collector portion of the collection card by allowing the blood to drip or squirt onto the card. Do not wipe the needle, ear or tail on the collection card. Discard the needle in an appropriate disposal container. 4. Before placing blood drops on blood cards, write an accurate customer name and animal ID in the spaces provided. Let the cards sit open and air dry before closing cover flap. Neogen: GeneSeek Operations 4131 N. 48th Street, Lincoln, NE igenity.support@neogen.com Neogen Corporation, Neogen and Igenity are registered trademarks of Neogen Corporation, Lansing, Michigan. AG

5 EQUIPMENT NEEDED To take a tissue sample using the Allflex Tissue Sampling Unit, the following tools are needed: 1. An Allflex tissue applicator 2. Allflex tissue punches Animals should be restrained during sampling (For example, by crowding in an alley or by using a head gate). TSU s should be stored at room temperature prior to use (for a maximum period of 12 months). Care should be taken not to expose the unused product to extremes of heat or cold prior to sampling. HOW TO USE THE ALLFLEX TISSUE APPLICATOR A. LOADING THE ALLFLEX TISSUE APPLICATOR 2 Ensure the tube retainer at the base of the applicator gun is open. Push the retainer clip to open. 1 Remove a TSU Punch from the packaging. Punches should be assembled as one piece: if they have come apart, reassemble. 3 Insert the punch into the tissue applicator as shown below. Release clip to lock punch into the device. 4 Carefully squeeze the applicator handles together, guiding the punch tip into place if necessary. When fully seated the gun bolt will rest flush against the red plastic clip. 5 Release the handle 6 Remove the red plastic clip by pulling it outward. TAKE CARE NOT TO CUT FINGER ON THE METAL CUTTER-IT IS VERY SHARP. 7 The applicator is now loaded and ready to take a sample. All parts are for one-off use only; do not re-use metal cutters.

6 B. TAKING THE SAMPLE 1 Ensure the tissue applicator is loaded with an unused tissue punch (the red plunger is visible in used punches). 2 Slide the gun over the ear, and position the cutter approximately 1 inch from the edge of the ear, taking care to avoid any obvious veins and ridges. Un-used punch Used punch 3 Sqeeze handles together to take a sample and then release to free the ear. Try to do this in one swift, fluid, motion. Move with the animal and don t fight its movements. Extract sample from this area. (1 inch from head, 2 inches from edge of ear) 4 Remove the punch from the device and check that sampling has been successful. If not, discard the sample and re-sample with a new punch. 5 Remove the used cutter from the applicator by pulling the handles apart. This will loosen the cutter. Discard safely. CAUTION: THE CUTTER IS VERY SHARP. Once the tissue sample has been extracted, care should be taken to store the product at room temperature if it will be dispatched to the laboratory within 2 weeks; otherwise, it is recommended that the samples are stored in a refrigerator or freezer to ensure complete preservation of the tissue sample. (The best results are obtained when the samples are analyzed within one year after the sample collection.) ALLFLEX USA, INC E. 14th Street Dallas Ft. Worth Airport, Texas (800)989-TAGS (8247) ALLFLEX CANADA 4135 Berard St-Hyacinthe, Québec J2S 8Z8 (866)505-TAGS (8247) PCT Patent Application WO 2010/066475

7 FREQUENTLY ASKED QUESTIONS 1. What do I need to do if one of my DNA Test Samples receives a fail from the Neogen Lab? a. The Owner/Breeder must order and complete payment for a new DNA Test and collect a new Test Sample. A failing test sample can be caused by contamination (dirt, hair, cleaning agents, tattoo ink/dye, etc.) or improper storage of the test sample. 2. Do I need to retest all animals submitted previously to UC Davis for DNA Testing? a. No, all genotype data previously tested through the TLBAA at UC Davis will transfer over. Progeny will have to be tested on STR for Parent Verification. 3. What if the Sire or Dam are deceased? a. If the Sire or Dam are deceased and a DNA Sample cannot be taken, a Letter of Evidence must be submitted to the TLBAA Office. The letter will be forwarded to the Board of Directors for approval. Please include all necessary information/photographs to show that the deceased party is the Sire/Dam to the calf. 4. Why do Hair Card Test Samples have a $4.00 additional fee? a. All Hair samples must be processed strand by strand. Individual hair follicles are separated from the hair strand before DNA can be extracted, this process is delicate, time consuming and must be done carefully. 5. What if Test Samples are mailed to Neogen before I receive my Order Number from the TLBAA Office? a. All samples that arrive at the Neogen Lab without an Order Number will be placed in the unidentified sample storage, until they can be properly identified. This will cause a delay in all DNA results. 6. How long will it take to get results from a Parent Verification DNA Test? a. Parent Verification results may take 3-4 weeks for completion, all results will be provided when they become available. 7. Why did my Tissue Sample fail at the Neogen Lab? a. All Tissue Sample Units contain a liquid that acts as a buffer between the air and tissue sample. This liquid must touch the Tissue Sample at all times, if the tissue sample is stuck in the top of the sample tube and the lid is securely sealed, invert the tube so that the tissue sample is in the buffer liquid. 8. I collected my test samples but have not received my Order Number from the TLBAA, how do I store my test samples? a. Hair Cards can be stored at room temperature in a reduced traffic area. b. Blood Cards must dry open to the air for 24 hours before they are shipped, do not freeze blood cards, keep them at room temperature in a reduced traffic area. c. Tissue Samples must be touching the buffer liquid (Question 7), tissue samples not sent within 24 hours of taking the sample can be refrigerated or kept at room temp. It is not required to ship the tissue sample with cold packs. TEST SAMPLES MUST BE MAILED TO NEOGEN. DO NOT SEND TEST SAMPLES TO TLBAA. ALL TESTING SAMPLE SUPPLIES MUST BE PURCHASED THROUH

8 Common Reasons for Samples that Fail DNA Testing General Issues 1. Biological Contamination a. Cross contamination between samples/animals b. Fecal matter and dirt in sample any foreign material in the sample may interfere with genotyping 2. Chemical Contamination a. Dye/Pigment from animal markers b. Insect repellant c. Cleaning agents 3. Improper Storage a. Heat exposure including leaving samples in vehicle or in hot sun. b. Exposure to foreign material including mold c. Improper frozen storage - the freeze/thaw cycle of a self-defrosting unit can degrade DNA d. Extended sample storage DNA degrades over time 4. Insufficient sample 5. Improper shipping Concerns Specific to Sample Type Hair cards 1. Too few or no follicles (<30) DNA only occurs in the root of the hair. The actual strands do not contain DNA. 2. Small follicles taken from young calves - hair samples should not be taken from calves < 6 months of age. Blood cards 1. Insufficient blood spotted on card - ensure the circle is filled and the blood is visible on both sides of the paper. 2. When spotting cards from stored blood, make sure to invert the tubes 4-5 times before spotting blood settles/separates over time. 3. Do NOT speed-dry the card using a hair-dryer as heat may damage the DNA. Whole blood 1. Wrong type of blood tubes are used. Use only purple-top EDTA tubes, these prevent clotting. 2. Store the blood in a refrigerator until shipped. Page 1 of 2

9 AllFlex Tags, Typifix Tags, and Tissue samples 1. No sample in tag 2. Sample is trapped in cap/does not enter tube sample will not be preserved 3. AllFlex tags and Typifix tags should be kept at room temperature to prevent DNA degradation. 4. Long term storage a. AllFlex Tags are viable for 12 months after collection. b. Tissue samples should be stored in the freezer. 5. Unpreserved tissue samples should be shipped next-day and never over a weekend or holiday. Semen 1. Store any semen samples at room temperature or a refrigerator. 2. Semen straws can easily be damaged in shipping. Proper packaging is especially important an empty ball point pen tube is recommended packaging

10 STR DNA TESTING WITH NEOGEN ALL TESTING SAMPLE SUPPLIES MUST BE PURCHASED BEFORE DNA TESTING CAN BEGIN 3 WAYS TO GET RESULTS: SNP GGP50K Short Tandem Repeat Analysis Single-nucleotide Polymorphism GeneSeek Genetic Profiler Also called snip testing Benefits of STR Testing Benefits of SNP Testing Benefits of GGP50 Testing ü Can be used for Parent Verification ü Analyzes Repeating DNA Markers ü Used on case-by case basis ü Accepted DNA Test ü Can be analyzed against other STR DNA Tests ü Can be bundled with a SNP DNA Test ü Can be used for Parent Verification ü Analyzes DNA Markers ü Used more regularly ü Widely accepted DNA Test ü Can be analyzed against other SNP DNA Tests ü Can be bundled with a STR DNA Test ü SNP ONLY; Can be used for Parent Verification ü Analyzes 50,000 DNA Markers ü Utilizes SNP Testing ü In Herd EPD s (Expected Progeny Differences) ü Widely accepted SNP DNA Test ü Can be analyzed against other SNP DNA Tests If you want to complete a Parent Verification DNA Test: All three animals must be tested by the same process. STR STR SNP SNP -STR can only be compared to STR- -SNP can only be compared to SNP- FREQUENTLY ASKED QUESTIONS: 1. What do I need to do if one of my DNA Test Samples receives a fail from the Neogen Lab? a.the Owner/Breeder must order and complete payment for a new DNA Test and collect a new Test Sample. A failing test sample can be caused by contamination (dirt, hair, cleaning agents, ink/dye, etc.) or improper storage of the test sample. 2. Do I need to retest all animals submitted previously to UC Davis for DNA Testing? a.no, all genotype data previously tested through the TLBAA at UC Davis will transfer over. Progeny will have to be tested on STR for Parent Verification. TEST SAMPLES MUST BE MAILED TO NEOGEN. DO NOT SEND TEST SAMPLES TO TLBAA. WHY BUNDLE STR AND SNP? SNP became the preferred method of Genotyping in A majority of cattle DNA tested through the TLBAA were tested on the STR method through UC Davis. STR is still reliable and available in most labs, but SNP is the future. Testing progeny with a bundle (STR and SNP) allows Parent Verification to the progeny without the need to retest Dams/Sires. The Progeny would be the first in a Pedigree to transition to SNP Testing. ALL TESTING SAMPLE SUPPLIES MUST BE PURCHASED THROUH

11 DNA TESTING PRICE LIST PRICE PER ANIMAL TESTED Ø SNP for parent verification...$15.00 Ø STR for parent verification...$15.00 Ø BUNDLE (STR & SNP) for parent verification...$25.00 Ø GGP50K +IGENTITY with parent verification...$ Includes one SNP DNA Test -In herd EPD capability SAMPLE TEST TYPES: Ø Blood Cards...($0.50 per card/10 card minimum order) Ø Hair Cards...($1.00 per card/10 card minimum order) Ø Tissue Sampling Unit...(21.00 for 10-unit pack/ Applicator Gun $45.00 each) Additional Fees to be expected: Ø $4.00 fee on ALL Hair Card Tests Ø Shipping and Handling TLBAA PO BOX 4430 Fort Worth, TX OFFICE: (817) Testing Samples MUST be sent with an ORDER NUMBER provided by the TLBAA Office. Samples sent in without an order number may not be identified properly and may result in increased wait time. TEST SAMPLES MUST BE MAILED TO NEOGEN. DO NOT SEND TEST SAMPLES TO TLBAA. ALL TESTING SAMPLE SUPPLIES MUST BE PURCHASED THROUH

12 REQUEST FOR DNA MARKERS (ONLY) If Animal is not registered, it MUST be recorded with the TLBAA Registration Office Name: Membership # Address: Phone: Name and Registration Number (if registered) of Animal to be DNA TESTED: Name: Registration # Male: Female: Date of Birth: Private Herd Number: Color: Name and Registration Number (if registered) of Animal to be DNA TESTED: Name: Registration # Male: Female: Date of Birth: Private Herd Number: Color: Name and Registration Number (if registered) of Animal to be DNA TESTED: Name: Registration # Male: Female: Date of Birth: Private Herd Number: Color: Return this form to: TLBAA PO BOX 4430 Fort Worth, TX OFFICE: (817) Testing Samples MUST be sent to Neogen with an ORDER NUMBER provided by the TLBAA Office. Samples sent in without an order number may not be identified properly and may result in increased wait time. TEST SAMPLES MUST BE MAILED TO NEOGEN. DO NOT SEND TEST SAMPLES TO TLBAA. ALL TESTING SAMPLE SUPPLIES MUST BE PURCHASED THROUH

13 1 REQUEST FOR PARENT VERIFICATION DNA TEST If Animal is not registered, it MUST be recorded with the TLBAA Registration Office Name: Membership # Address: Phone: Name and Registration Number (if registered) of Animal Parentage Being Verified: Name: Registration # Male: Female: Date of Birth: Private Herd Number: Color: SIRE(S) IN QUESTION Sire #1 Name: Registration # Sire #2 Name: Registration # Dam #1 DAM(S) IN QUESTION Name: Registration # Dam #2 Name: Registration # Return this form to: TLBAA PO BOX 4430 Fort Worth, TX OFFICE: (817) Testing Samples MUST be sent to Neogen with an ORDER NUMBER provided by the TLBAA Office. Samples sent in without an order number may not be identified properly and may result in increased wait time. TEST SAMPLES MUST BE MAILED TO NEOGEN. DO NOT SEND TEST SAMPLES TO TLBAA. ALL TESTING SAMPLE SUPPLIES MUST BE PURCHASED THROUH

14 REQUEST FOR GGP50K TESTING (with parent verification) If Animal is not registered, it MUST be recorded with the TLBAA Registration Office Name: Membership # Address: Phone: Name and Registration Number (if registered) of Animal Parentage Being Verified: Name: Registration # Male: Female: Date of Birth: Private Herd Number: Color: GGP50K SIRE(S) IN QUESTION Sire #1 Name: Registration # GGP50K Sire #2 Name: Registration # GGP50K DAM(S) IN QUESTION Dam #1 Name: Registration # GGP50K Dam #2 Name: Registration # GGP50K Return this form to: TLBAA PO BOX 4430 Fort Worth, TX Testing Samples MUST be sent to Neogen with an ORDER NUMBER provided by the TLBAA Office. Samples sent in without an order number may not be identified properly and may result in increased wait time. TEST SAMPLES MUST BE MAILED TO NEOGEN. DO NOT SEND TEST SAMPLES TO TLBAA. ALL TESTING SAMPLE SUPPLIES MUST BE PURCHASED THROUH