Frequently Asked Questions
Q1: How do I order transgenic fly services from GenetiVision?
Q2: How should DNAs be prepared for injection?
Q3: What type of guarantee does GenetiVision provide for its services?
Q4: How many independent transgenic flies should I expect?
Q5: Do you accept credit cards for payment?
Q6: What is the turn around time for GenetiVision services?
Q7: What are the prices for GenetiVision services?
Q8: How do I screen for transgenic flies after I receive injected larvae?
Q9: How do I order BAC duplication stocks?
Q1: How do I order transgenic fly services from GenetiVision?
Simply click here. A Word document will download to your computer. Fill out the form, including a valid FedEx account number that we will use to send your larvae or flies to you, and a Purchase Order (PO) number. If you intend to use a credit card for payment, please do not include your card information on the Order Form. Instead, please write "Credit Card" in the space provided for a PO number at the bottom of the Order Form. If you choose to pay by credit card, you will receive an invoice via PayPal when your work is complete and you will enter your credit card information only at the PayPal website. Please email the completed form to us as a Word ".doc" file (please do not save as a PDF or as ".docx"). In addition, please send a hard copy of the same completed form along with your DNA samples to:
GenetiVision Corporation
3844 Greenbriar Drive
Stafford, TX 77477
Q2: How should DNAs be prepared for injection?
We find that the DNA preparation step is very important for successful transgenesis. We recommend using either the Qiagen or Invitrogen Midi-prep kits for DNA preparation. In no case should Mini-prep DNA be sent to us as this is toxic to embryos. All DNA samples should be provided in EB (10 mM Tris-Cl, pH 8.5) or water at 500 ng/μl. We request that you send us 20 µl of such DNA. For larger constructs (>25 kb), 30 µl of 1 µg/µl DNA is required. To minimize the chances of having to resend DNA samples, for construct greater than 25 kb, it is highly recommended that you filter the DNA before sending to us. To filter the DNA, we use the Pall Nanosep® MF Centrifugal Device 0.2 µM. Please make sure that the DNA concentration is at least 500 ng/µl after filtering.
Q3: What type of guarantee does GenetiVision provide for its services?
We guarantee all PhiC31 injections for DNAs less than 40 kb in length and all P-element injections for DNAs less than 20 kb in length (including vector sequences). This means that if you do not find any transgenics with our first round of injections, we will do another round for free right away. If you still do not get any transgenics (this is extremely rare), then we normally will not do another round of injections as it is most likely a problem with the construct or the DNA prep. If it is clear that we have made a mistake (also extremely rare!), then we would inject again for free. We cannot offer a guarantee for DNAs greater than 40 kb in length since these are relatively low probability events that are very sensitive to very small changes in conditions. Nevertheless, we have found that most of our injections for large DNAs are in fact successful, including hundreds of DNAs >100 kb. Due to the difficulties associated with large DNA injection, we require that we prepare your DNA for constructs >40 kb.
Q4: How many independent transgenic flies should I expect?
3-5 independent transgenic flies are normally obtained for P-element or Phi-C31 injections of DNAs less than 20 kb, often many more. We guarantee three independent transgenic flies for P-element injections and at least one independent insertion for PhiC31 (since that is all you should need).
Q5: Do you accept credit cards for payment?
Yes, we accept all major credit cards. However, a 3% service charge will be applied to cover the processing fee charged to us by PayPal (4% for orders from outside the USA). If you choose to pay by credit card, please do not include your card information on the Order Form. Instead, please write "Credit Card" in the space provided for a PO number at the bottom of the Order Form. When your work is complete you will receive an invoice via e-mail from PayPal and you will enter your credit card information only at the PayPal website.
Q6: What is the turn around time for GenetiVision services?
Once we receive your DNA, normally we will send out G0 larvae within 7 to 10 business days. Add two to three days if we are also preparing your DNA. Add four to five weeks if we are selecting transgenic flies for you (two generations = 2x two weeks: one generation for the G0 larvae to become adults and another generation to obtain F1 progeny of G0 adults which will be screened for transgenic insertions).
Q7: What are the prices for GenetiVision services?
We provide remarkably affordable prices starting at just $220 for standard P-element or PhiC31 site-specific transgenesis.
For complete pricing details for all of our services, please click here.
Q8: How do I screen for transgenic flies after I receive injected larvae?
We are currently using two basic methods for creating transgenic flies.
P-element Transgenesis Using a white-plus DNA for Injection
For P-element mediated transgenesis, we inject your white-plus DNA into embryos derived from a cross of y w males to y w ; Ki delta-2-3 virgins. Ki is a dominant mutation on the third chromosome which produces kinked (i.e., shortened and twisted) bristles. Therefore, 100% of the G0 flies derived from the larvae we send to you will be white-minus (since the white-plus gene in your vector will not be expressed in anterior structures until the F1) and heterozygous for the Ki delta 2-3 third chromosome (please note that the phenotype from Ki is not fully penetrant, it is possible that flies will not show kinked bristles). In contrast, 50% of F1 progeny (where you look for white-plus transgenic animals) will possess various combinations of these markers depending on the sex of the G0 fly. For crosses with G0 males, Ki and delta-2-3 will remain linked (since there is no recombination in males) and one can select against Ki to rapidly select against delta-2-3, particularly in the case of very weak white-plus insertions where white-plus eye color mosaicism may be difficult to score. However, the Ki delta 2-3 chromosome is not a balancer and Ki and delta-2-3 are essentially unlinked (Ki is at 83DE and delta-2-3 is at 99B). Therefore, if you mate a G0 female, Ki and delta-2-3 will often be separated by recombination and thus Ki cannot be used to select for flies which no longer contain delta-2-3. In this case, it is best to select against mosaic white-plus eyes. It is necessary to remove delta-2-3 from your stock in order to establish and maintain a stable P-element insertion. In rare cases it may be that all your white-plus F1 flies are mosaic. In such cases, just outcross these to white-minus flies again (as you did with your G0 flies) and look for white-plus non-mosaic animals in the F2.
PhiC31 Mediated Transgenesis Using a white-plus DNA for Injection
In the PhiC31 site-specific integration system, we inject DNA into embryos containing any docking site of your choice. We are currently offering the docking sites at no additional charge:
ZH-2A: (X) 2A3
attP18: (X) 6C12
su(Hw)attP8: (X) 8E10
VK37: (2L) 22A3
JK22C: (2L) 22C3
attP40: (2L) 25C6
JB38F: (2L) 38F1
su(Hw)attP5: (2R) 50F1
ZH-51C: (2R) 51C1
attP1: (2R) 55C4
VK22: (2R) 57F5
VK01: (2R) 59D3
VK31: (3L) 62E1
VK33: (3L) 65B2
JK65C: (3L) 65C3
JK66B: (3L) 66B4
attP2: (3L) 68A4
JK73A: (3L) 73A9
VK05: (3L) 75A10
ZH-86Fb: (3R) 86F8
VK40: (3R) 87B10
su(Hw)attP1: (3R) 87B13
VK27: (3R) 89E11
VK20: (3R) 99F8
We normally inject embryos that are homozygous for both an X-chromosome source of PhiC31 integrase and one of the docking sites listed above. If you wish to use a different docking site you will need to supply the flies, wait 1-2 generations for these flies to be expanded, and pay a $50 surcharge. G0 animals from our normal injections will be homozygous for the integrase-expressing X chromosome. While it is best to remove this chromosome from your transgenic stock, the insertion is stable in the continued presence of integrase since the integration event occurs between attB (in the injected DNA) and attP (in the docking chromosome) sites, generating attR and attL sites which are no longer substrates for the integrase. As in P-element transgenesis, G0 flies will not express the white-plus gene and you must wait until the F1 to select transgenic flies.
Q9: How do I order BAC Duplication stocks?
Simply click here. A Word document will download to your computer. Fill out the form, including a valid FedEx account number that we will use to send your flies to you, and a Purchase Order (PO) number. If you intend to use a credit card for payment, please do not include your card information on the Order Form. Instead, please write "Credit Card" in the space provided for a PO number at the bottom of the Order Form. If you choose to pay by credit card, you will receive an invoice via PayPal when we ship you stocks and you will enter your credit card information only at the PayPal website. Please email the completed form to us as a Word ".doc" file (please do not save as a PDF or as ".docx").