Stratified haploid identification system through the R1-nj kernel and reduced seedling vigor in tropical maize germplasm




Abstract. Thawarorit A, Dermail A, Lertrat K, Chankaew S, Suriharn K. 2023. Stratified haploid identification system through the R1-nj kernel and reduced seedling vigor in tropical maize germplasmBiodiversitas 24: 4262-4268. Haploid identification through the R1-nj marker is commonly applied in in-vivo doubled haploid technology in maize, although misclassification issues often occur. We explored the innate properties of maternal haploids at the seedling stage to verify true haploids and to reduce false positives. There are many questions about whether reduced seedling vigor is effective for haploid verification in tropical maize backgrounds. This study aimed to evaluate the haploid induction rate of Stock6-derived Fpopulation inducers and to investigate the effectiveness of reduced seedling vigor at the V2/V3stage as an alternative marker to verify putative haploids. Field trials and haploid induction were conducted in the rainy season of 2021 and the dry season of 2021/22 in Khon Kaen, Thailand. Two tropical source germplasm: P789 and S7328, were included as donor females. Haploids were identified through the R1-nj marker and reduced seedling vigor. Significant reduction of haploid frequency up to 2.5% on average between the two identification methods, indicating that a considerable number of false positives could be found using reduced seedling vigor. Inducer population K11 was promising since it performed a stable ability to produce haploids over donors and seasons and had a low misclassification rate. Further breeding strategies and haploid selection schemes are discussed.


Ao GM, Zhao SX, Li GH. 1982. In vitro induction of haploid plantlets from unpollinated ovaries of corn (Zea mays L.). Acta Genetica Sinica 9(4): 281–283.
Baleroni AG, Ré F, Pelozo A, Kamphorst SH, Carneiro JWP, Rossi RM, Scapim CA. 2021. Identification of haploids and diploids in maize using seedling traits and flow cytometry. Crop Breeding and Applied Biotechnology 21(4): 1-8.
Chaikam V, Martinez L, Melchinger AE, Schipprack W, Boddupalli PM. 2016. Development and validation of red root marker?based haploid inducers in maize. Crop Science 56(4): 1678-1688.
Chaikam V, Molenaar W, Melchinger AE, Boddupalli PM. 2019. Doubled haploid technology for line development in maize: technical advances and prospects. Theoretical and Applied Genetics 132: 3227-3243.
Chaikam V, Nair SK, Babu R, Martinez L, Tejomurtula J, Boddupalli PM. 2015. Analysis of effectiveness of R1-nj anthocyanin marker for in vivo haploid identification in maize and molecular markers for predicting the inhibition of R1-nj expression. Theoretical and Applied Genetics 128: 159-171.
Chaikam V, Nair SK, Martinez L, Lopez LA, Utz HF, Melchinger AE, Boddupalli PM. 2018. Marker-assisted breeding of improved maternal haploid inducers in maize for the tropical/subtropical regions. Frontiers in Plant Science 9: 1527.
Coe Jr EH. 1959. A line of maize with high haploid frequency. The American Naturalist 93(873): 381-382.
Department of Agriculture, Thailand. 2022. Thai Agricultural Practice.
Dermail A, Chankaew S, Lertrat K, Lübberstedt T, Suriharn K. 2021. Selection gain of maize haploid inducers for the tropical savanna environments. Plants 10(12): 2812.
Eder J, Chalyk S. 2002. In vivo haploid induction in maize. Theoretical and Applied Genetics 104: 703-708.
Erenstein O, Jaleta M, Sonder K, Mottaleb K, Prasanna BM. 2022. Global maize production, consumption and trade: trends and R&D implications. Food Security 14(5): 1295-1319.
Geiger HH, Gordillo GA. 2009. Doubled haploids in hybrid maize breeding. Maydica 54(4): 485-499.
Geiger HH. 2009. Doubled haploids. In Bennetzen JL, Hake S (eds) Maize Handbook - Volume II: Genetics and Genomics. Springer Science and Business Media, New York.
Gilles LM, Khaled A, Laffaire JB, Chaignon S, Gendrot G, Laplaige J, et al. 2017. Loss of pollen?specific phospholipase NOT LIKE DAD triggers gynogenesis in maize. The EMBO Journal 36(6): 707-717.
Gomez KA, Gomez AA. 1984. Statistical Procedure for Agricultural Research. John Wiley and Sons, Singapore.
Greenblatt IM, Bock M. 1967. A commercially desirable procedure for detection of monoploids in maize. Journal of Heredity 58(1): 9-13.
Kelliher T, Starr D, Richbourg L, Chintamanani S, Delzer B, Nuccio ML, et al. 2017. MATRILINEAL, a sperm-specific phospholipase, triggers maize haploid induction. Nature 542(7639): 105-109.
Liu Z, Wang Y, Ren J, Mei M, Frei UK, Trampe B, Lübberstedt T. 2016. Maize doubled haploids. Plant Breeding Reviews 40: 123-166.
Melchinger AE, Schipprack W, Friedrich Utz H, Mirdita V. 2014. In vivo haploid induction in maize: identification of haploid seeds by their oil content. Crop Science 54(4): 1497-1504.
Melchinger AE, Schipprack W, Würschum T, Chen S, Technow F. 2013. Rapid and accurate identification of in vivo-induced haploid seeds based on oil content in maize. Scientific Reports 3(1): 2129.
Nanda DK, Chase SS. 1966. An embryo marker for detecting monoploids of maize (Zea mays L.). Crop Science 6(2): 213-215.
Prasanna, BM. 2012. Doubled haploid (DH) technology in maize breeding: An overview. In Prasanna BM, Chaikam V, Mahuku G (eds) Doubled Haploid Technology in Maize Breeding: Theory and Practice. CIMMYT, Mexico.
Prigge V, Sánchez C, Dhillon BS, Schipprack W, Araus JL, Bänziger M, Melchinger AE. 2011. Doubled haploids in tropical maize: I. Effects of inducers and source germplasm on in vivo haploid induction rates. Crop Science 51(4): 1498-1506.
Rotarenco V, Dicu G, Fuia S. 2010. New inducers of maternal haploids in maize. Maize Genetics Cooperation Newsletter 84: 21-22.
Sekiya A, Pestana JK, Silva MGBD, Krause MD, Silva CRMD, Ferreira, JM. 2020. Haploid induction in tropical supersweet corn and ploidy determination at the seedling stage. Pesquisa Agropecuária Brasileira 55: 1-10.
Sintanaparadee P, Dermail A, Lübberstedt T, Lertrat K, Chankaew S, Ruanjaichon V, Phakamas N, Suriharn K. 2022. Seasonal variation of tropical savanna altered agronomic adaptation of Stock-6-derived inducer lines. Plants 11(21): 2902.
Statistix R. 2013. Statistix 10 Analytical Software. Tallahassee, Florida.
Uliana Trentin H, Batîru G, Frei UK, Dutta S, Lübberstedt T. 2022. Investigating the effect of the interaction of maize inducer and donor backgrounds on haploid induction rates. Plants 11(12): 1527.
Zhong YU, Liu C, Qi X, Jiao Y, Wang D, Wang Y, et al. 2019. Mutation of ZmDMP enhances haploid induction in maize. Nature Plants 5(6): 575-580.

Most read articles by the same author(s)