Frequently Asked Questions about Wheat TILLING

1. How do I germinate wheat seeds?
   Upon receiving seeds, we recommend first germinating wheat seeds in moistented germination paper, either wrapped as a tight cylindar or in the base of a petri dish. Seeds should then be refridgerated at 4 degrees C for 3-5 days before moving to room temperature. After approximately one week, seedlings should be ready to transfer to soil.   
    
2. What do I get when I order a seed stock?
   For each mutant line, approximately 10-40 M4 seeds will be dispatched, depending on the availability of the seed stocks of the required line. We will also include approximately 20 seeds of unmutagenized wild-type Kronos to carry out backcrossing.
    
3. Where do I look for phenotypes?
   Be aware that M4 mutant lines carry a high density of EMS-induced mutations (Roughly one mutation per 38.4kb). This can adversely affect germination and result in abnormal and highly variable growth and development. Therefore, to reliably assess the phenotypic effects of the mutation, we recommend performing at least one and preferably 2-4 backcrosses to an unmutangenized Kronos parent to reduce the mutation load. 
    
4. I get mutants in my BLAST results, but when I click on the visualization, the mutation track does not load.  How do I fix this?
   Users should clear their internet browser cache or use private browsing mode.
   HowTo for Chrome  •  HowTo for Firefox
    
5. In the mutation effect prediction, what is the difference between splice_region_variant and splice_acceptor_variant/splice_donor_variant?
   These are formal definitions as defined by the Sequence Ontology.  Splice_acceptor/Splice_donor variants only occur at the 2 base pairs flanking an exon. Splice_region_variants can occur 3-8 base pairs of the intron border, or 1-3 base pairs of the exon border.  To find out more about these definitions, read the formal VEP definitions here.
6. I have received a seed but am having trouble confirming the mutation. What is going on?
   The confidence intervals of high (99%), medium (98%), and low (95%) are probabilities that we have correctly identified a true mutation. This is different than the probability the mutation still exists within the seed stocks. Since this resource was created using M2 plants, but we are distributing bulked M4 seeds, if the mutation was heterozgyous in the M2, there is a chance that the mutation could have been lost through segregation when advancing the generations.