CUTTER CMS, LLC

ACQUIRED BY GRO ALLIANCE

Gary L. Cutter, Ph.D.

Corn Cytoplasmic Male Sterility Conversions Made Genetically Pure in 2-3 Generations

(Web site information updated 11/24/2018)

Since October 2011, Cutter CMS, LLC has provided a service to the seed corn industry of developing cytoplasmic male sterility conversions of corn inbred lines used as females in seed production by using a unique efficient technique. The use of such male sterile female parents significantly reduces the costs of seed production by (1) reducing or eliminating the costs of detasseling, (2) increasing seed yields from 15 to 30% as compared to mechanically detasseled seed production, (3) reducing the risks of insufficient detasseling labor and weather influences, and (4) decreasing the exposure to safety incidents due to detasseling by hand.

Since beginning the service 7 years ago, a total of 154 projects (inbred/cytoplasm combinations) have been entertained from 11 clients.

In June 2018, Gro Alliance, a seed corn and soybean production company with seed corn nursery services, acquired the assets and intellectual property of Cutter CMS, LLC and began the process with new clients. (See the press release at this link).The agreement allows Cutter CMS, LLC to continue work with its present clients until 2023, after which these clients will be served by Gro Alliance. A two year training of the process is being provided followed by availability for consultation for subsequent years. Any new inquiries will be handled by Jim Schweigert of Gro Alliance at jim.schweigert@groalliance.com.

Technique Used:

The conversion technique used is the “indeterminate gametophyte (ig) system” involving paternal haploids and cytoplasmic male sterility. Significant advantages of the results of this conversion technique include:

• The resulting conversions from this method will have 100% of the nuclear genetic component of the converted inbred in a male sterile cytoplasm (C or SD).
  
• Such male sterile females will cause no hybrid yield drag that is possible due to remnant linkages that will result from traditional backcross conversion methods, even with genetic markers yielding 95 to 99% recovery. This is especially important for specialty corns such as popcorn or sweet corn. No yield testing is necessary.
• Inbreds are converted in only two breeding generations. An additional generation is used for phenotypic verification and the initial hand pollinated seed increase to produce an amount sufficient for breeder seed production.

Experience of Gary Cutter, Ph.D., Owner and Corn Breeder of Cutter CMS, LLC:

Gary retired in 2011 with over 36 years of experience in the seed corn industry with a major seed company in the areas of corn breeding, research station and regional management, corn seed production research, male sterility conversions and parent seed management. He had been involved with the “ig” male sterility conversion system since the mid 1970’s when Dr. John Laughnan provided the initial genetic stocks to the industry corn breeders during the Illinois Corn Breeders School. There have been inbred conversions from his development programs that have been used significantly in the industry during the past 20 years.

Gary earned a Ph.D. in Plant Breeding and Genetics at the University of Wisconsin-Madison after earning BS and MS degrees in Ag Education and Agronomy, respectively, at Purdue University. He continues to be a student of the “ig” technique and has been developing efficiency improvements to the system. Gary started this business after his corporate retirement because he enjoys the outdoor field nursery work, seeing the value added results of an inbred male sterile conversion going through the paternal haploid stage and adding value as a perfect genetic and phenotypic recovery without using molecular methods.

How the “ig” Male Sterile Conversion System Works:

The system utilizes the effects of the “ig1” gene on the abnormal development of the female gametophyte in the W23 genetic background. Progeny of a cross of an inbred to be converted as a pollinator on the ig stock will include a low percentage of haploid plants, about 2/3 of which are of paternal origin genetically. The stock used as the female in the cross includes the male sterile cytoplasm which will be inherited maternally by the paternal haploids; thus these paternal haploid plants will carry 100% of the pollen donor’s nuclear genome along with the male sterile cytoplasm. These haploid plants are then backcrossed as the female to the donor inbred and due to chance a few kernels are generally produced. Nearly all these kernels originate from a few “unreduced female gametes” that occur by chance during meiosis in the haploid plant. Nearly all of these kernels are viable and grow as diploid plants and are identical phenotypically and genetically to the donor inbred with the desired exception of being male sterile due to the cytoplasm genetic effects if the donor inbred does not carry restoring genes. These are the plants that are increased during the 3rd generation of the process, where the phenotype and male sterility is verified. Of course the increase of the male sterile plants is done by crossing with the original inbred being used as the “maintainer”.

Service History and Success Rate (as of 11/23/2018):

Since beginning the service in 2011, a total of 154 projects (inbred/cytoplasm combinations) have been completed or are in progress from 11 clients. A total of 33 of these projects are presently active, undergoing either the first, second or third generation of the process. Clients have the option of dropping a project before the planting date. Of those projects that have not been dropped, 97% have been completed; 81% were completed during the 3 generations planned. Delays of completion have been primarily due to no paternal haploids being produced or failure to set viable seed on the paternal haploids due to silking, insect or disease problems.

Options, Timelines and Fees of the “ig” CMS Conversion Service

The conversion process may be started either in the off-season or summer nursery. However, the second generation of the process must be done during the home summer nursery because of the attention to detail necessary in identification, pollination and care of the paternal haploid plants. If the process is begun in the off-season nursery, then it can be finished in three consecutive growing seasons (off-season, home, and off-season) with seed of the completed conversion provided for a spring planted increase by the client. If the first generation of the project is done during the summer, the second generation will still be done the following summer. This would result in two years to completion instead of 1 ½ years. As noted earlier, over 80% of the time seed of the conversion is generally provided after 3 generations. The remaining projects may need to have the second generation repeated due to lack of seed set on the paternal haploid plant or insect or disease damage to the seed set on the haploid plant.

Following is a description of each of the 3 generations of the process:

First Generation: For each inbred the client will provide the following by November 15 (or April 15 for summer nursery):

• Completion of your legal agreement if necessary to meet the deadline.
• Provide flowering comparisons of each inbred to facilitate a nick with my conversion stocks
• Provide inbred genetic background and descriptive characteristics (such as silk color and cob color) to facilitate the paternal haploid selection process
• A nonrefundable initiation fee will be invoiced for each inbred
• Provide a minimum of 200 kernels of viable seed of the inbred
• Seed will be planted with any delays necessary and plants crossed onto “ig” conversion stock(s) of both the C and SD cytoplasms.
• Testcrosses will also be made to inbred testers of these cytoplasms and grown out during the next generation to assess the inbred’s capability to be sterilized, having no residual restorer genes.

Second Generation:

• Any inbred may be discontinued by the client for any reason before planting this generation. Notification due dates are April 15 to avoid planting the second generation and to any avoid additional fees for continuing and finishing the conversion. A cytoplasm project will also be discontinued if the grow-outs show restoration of fertility in the male sterile cytoplasm tested, unless the client prefers to continue it.
• A nonrefundable continuation fee per inbred will be invoiced before planting in the summer nursery for each inbred/cytoplasm combination (project).
• Paternal haploid plants of the source inbred in the male sterile cytoplasm will be selected and backcrossed to the source inbred to produce some seed with unreduced female gametes that are expected to occur. Such seed generally results to be the completed conversion. A range of 0 to 40 seeds has previously been set on a paternal haploid plant generally depending on genetics. On occasion, complete seed set was noted to occur on spontaneously doubled paternal haploids, where such seed may be available a generation earlier. This has happened in approximately 5% of the projects. No colchicine or chromosome doubling agent is used.

Third Generation:

• Again, any project may be discontinued by the client for any reason before planting this generation. Notification due dates are September 1 to avoid planting the third generation and to any avoid additional fees for continuing and finishing the conversion.
• Seed from the backcrossed paternal haploids will be planted adjacent to the original inbred. The sterility and the plant phenotype will be verified by visual comparison. An increase of the converted inbred will be made expecting to produce from 250 up to 10,000 K depending on paternal haploid seed production. Increased seed of acceptable inbred conversions will be provided by April 1 after receiving the final payment. Seed may be returned on the ear if desired, or a few ears may be left unshelled for the client to see the ear type. Digital pictures of the ears can also be taken and made available if desired.
• The final fee payment will be invoiced prior to the return shipment of the seed. Shipping charges will also be assessed.

Summary of Fees: New conversion projects will be handled by Gro Alliance. Their total charge for the conversion of an inbred is $10,000, of which $2,500 is for initiation, $3,500 for the second generation planting, and $4000 before the return of seed increase after the third generation. This includes one or both cytoplasm conversions (C and SD) of the same inbred where it does not have residual restorer genes identified by the testcross grow-out. If the conversion is not completed due to the choice of the client or the presence of restorer genes, only the first and/or second generation charges are applied.

Additional Considerations:

• Overcoming the problem of not having hybrid yield data available in time to select inbreds to submit by November 15.
  
  Generally breeders rely on hybrid yield data to make the final decision for an inbred to advance to the next level to enter the conversion program and these data are not available until mid-November. The winter nursery will be in Chile and is desired to be planted near November 1. A late decision can be accommodated by receiving a larger group of inbreds that are being tested and planting them in the winter nursery at that time before final decision is made. Notification of the “go forward” selected inbreds by December 15 will allow the rows of the dropped inbreds to be cut out before pollination and only the selected inbreds will be crossed to the conversion stocks. The initial invoice will be for a targeted number of selected inbreds and can be adjusted during the second conversion generation. There will be no charge for those inbreds cut out and discarded from the program before pollination. This has been done with one client in the past with Cutter CMS, LLC.
  
  
• A client strategy to add traits to the non-traited cytoplasmic male sterile conversion
  
  Traits are generally added to a non-traited male sterile version by backcrossing two generations and selecting the homozygotes, either by testcrossing or molecular methods. In this case, only a few pollinations are needed and only a few converted male sterile plants are necessary. This leads to an option by the client to receive the seeds from the paternal haploid, if enough are available and backcross a generation earlier than the timeline described above. As these seeds are grown out for backcrossing, you may assess for male sterility and phenotype at the same time. The general protocol for a back-up increase will be followed in case a problem occurs and it also allows assessment of the sterility and phenotype in our nursery. Theoretically, if all goes well, a traited male sterile version can be made only 1 year after the original line is selected for such advance. In receiving seed at this time, one can expect to have from 5 to 100 kernels to work with. This seed would be provided to be grown ear row from each paternal haploid plant to allow for discarding any family that does not have the recurrent inbred characteristics for any unexpected reason (this has not been necessary in past experiences). Up to 10 seeds will be retained for a 3rd generation verification and increase in our 3rd generation nursery; this increased seed will also be provided to the client. Seed of the male sterile conversions from the paternal haploids may be provided before October 15 from the summer nursery and upon receipt of the final fee payment. This final payment would be refunded if the conversion is not correct according to both client’s and our nursery observations, which is expected to be a rare event based on past experience.
  
  
• Client testcrosses inbred onto male sterile cytoplasm before submitting for conversion. A client may determine from a testcross and grow-out he has done in advance the male sterile cytoplasm that would give the inbred effective sterility when converted. The grow-out can also be assessed during the first generation in crossing to the stocks. If in the relatively rare case the inbred is fertile in both cytoplasms, this would avoid taking an inbred through the 2nd generation where the fee of $3,500 to continue could be saved. If earlier testcrossing is desired as described, seed of B73CMS-C and B73CMS-SD can be provided to the client to use as a tester. The downside of this would be the loss of 1 or 2 growing seasons after the selection of inbreds to submit.
  
  

Contact Information:

Cutter CMS, LLC
Gary L. Cutter, Ph.D.
Cell phone: 1 (309) 825-5149
Email: gary@cuttercms.com

Gro Alliance Jim Schweigert
Cell Phone: 1 (608) 732-8459
Email: jim.schweigert@groalliance.com