Molecular Breeding

, 39:31 | Cite as

Marker-assisted backcross breeding for enhancing β-carotene of QPM inbreds

  • Deepti B. SagareEmail author
  • Prashant Shetti
  • M. Surender
  • S. S. Reddy


Maize possesses natural variations for crtRB1-carotene hydroxylase) and lcyE (lycopene-ε-cyclase) which significantly enhances provitamin A (proA) concentration. Enhancement of kernel proA carotenoids in QPM genetic background is of greater value. In this study, allelic variability for proA carotenoids was analyzed to identify promising β-carotene donor to introgress into QPM inbreds employing molecular breeding. Maize inbred MGU23379 (6.31 μg/g of β-carotene) was used as β-carotene donor to cross with recurrent parents (RPs), CB6-36 (CBML6) and CB7-28 (CBML7). In conversion program, F1, BC1F1, BC2F1, BC2F2, BC2F3, and BC2F4 materials were generated. In each generation, foreground selection was carried out with crtRB1-3′TE and umc1066. The crtRB1-3′TE segregated as per the expectation. Selection for modifiers, recombinants, and background genome of RPs was carried out in BC populations. The individuals with high recovery of recurrent parent genome were retained. Tryptophan/lysine content in introgressed progenies was on par, but β-carotene content was significantly high (6.25and 6.80 μg/g) compared to original inbreds (0.71 and 1.29 μg/g). Phenotypic data recorded for different traits in the BC2F3 populations did not show any significant difference between the converted BC2F3 families and their RPs. Also, grain yield of converted inbreds (PVCBML6 and PVCBML7) was on par with their original lines. In PVCBML6, stem anthocyanin pigmentation was reduced and silk color was changed to dark pink; whereas in PVCBML7, tassel was more erect and sparer; and silk color was changed to light pink compared to original inbreds. Converted inbreds provides an ideal platform for stacking number of nutritionally important traits.


QPM opqaue2 crtRB1-3′TE β-Carotene Provitamin A Recurrent parent genome Molecular breeding 



First author is thankful to Department of Science and Technology (DST), Govt. of India, for providing INSPIRE fellowship to pursue doctoral studies [IF120799]. All authors are thankful to Institute of Biotechnology and Maize Research Centre, PJTSAU, Hyderabad, for providing lab and field facilities to conduct the research.

Authors’ contributions

Conception and design of study: SSR and DBS, Development of segregating progenies and molecular analysis: DBS. Morphological characterization: DBS and PS. Phenotyping for kernel quality, biochemical analyses: DBS, MS, and PS. Statistical analyses: PS. Drafting of the manuscript: DBS and PS.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

11032_2019_939_MOESM1_ESM.pdf (72 kb)
ESM 1 (PDF 72 kb)
11032_2019_939_MOESM2_ESM.pdf (258 kb)
ESM 2 (PDF 257 kb)
11032_2019_939_MOESM3_ESM.pdf (177 kb)
ESM 3 (PDF 177 kb)
11032_2019_939_MOESM4_ESM.pdf (377 kb)
ESM 4 (PDF 377 kb)
11032_2019_939_MOESM5_ESM.pdf (281 kb)
ESM 5 (PDF 281 kb)
11032_2019_939_MOESM6_ESM.pdf (167 kb)
ESM 6 (PDF 166 kb)
11032_2019_939_MOESM7_ESM.pdf (160 kb)
ESM 7 (PDF 159 kb)
11032_2019_939_MOESM8_ESM.pdf (233 kb)
ESM 8 (PDF 233 kb)
11032_2019_939_MOESM9_ESM.pdf (163 kb)
ESM 9 (PDF 162 kb)
11032_2019_939_MOESM10_ESM.pdf (224 kb)
ESM 10 (PDF 223 kb)
11032_2019_939_MOESM11_ESM.pdf (225 kb)
ESM 11 (PDF 225 kb)
11032_2019_939_MOESM12_ESM.pdf (163 kb)
ESM 12 (PDF 163 kb)
11032_2019_939_MOESM13_ESM.pdf (296 kb)
ESM 13 (PDF 296 kb)


  1. Azmach G, Gedil M, Menkir A, Spillane C (2013) Marker-trait association analysis of functional gene markers for provitamin A levels across diverse tropical yellow maize inbred lines. BMC Plant Biol 13(1):227. CrossRefPubMedPubMedCentralGoogle Scholar
  2. Babu R, Prasanna BM (2014) Molecular breeding for quality protein maize (QPM). In: Tuberosa R, Frison E (eds) Genomics of plant genetic resources. Springer, Dordrecht, pp 489–505. CrossRefGoogle Scholar
  3. Babu R, Nair SK, Kumar A, Venkatesh S, Sekhar JC, Singh NN, Srinivasan G, Gupta HS (2005) Two-generation marker-aided backcrossing for rapid conversion of normal maize lines to quality protein maize (QPM). Theor Appl Genet 111(5):888–897. CrossRefPubMedGoogle Scholar
  4. Babu R, Rojas NP, Gao S, Yan J, Pixley K (2013) Validation of the effects of molecular marker polymorphisms in LcyE and CrtRB1 on provitamin A concentrations for 26 tropical maize populations. Theor Appl Genet 126(2):389–399. CrossRefPubMedGoogle Scholar
  5. Bain LE, Awah PK, Geraldine N, Kindong NP, Sigal Y, Bernard N, Tanjeko AT (2013) Malnutrition in Sub-Saharan Africa: burden, causes and prospects. Pan Afr Med J 15(1).
  6. Chander S, Guo YQ, Yang XH, Zhang J, Lu XQ, Yan JB, Song TM, Rocheford TR, Li JS (2008) Using molecular markers to identify two major loci controlling carotenoid contents in maize grain. Theor Appl Genet 116(2):223–233. CrossRefPubMedGoogle Scholar
  7. Danson J, Mbogori M, Kimani M, Lagat M, Kuria A, Diallo A (2006) Marker-assisted introgression of opaque2 gene into herbicide tolerant elite maize inbred lines. Afr J Biotechnol 5:2417–2422 Available at: Google Scholar
  8. Feng F, Wang Q, Liang C, Yang R, Li X (2015) Enhancement of tocopherols in sweet corn by marker-assisted backcrossing of ZmVTE4. Euphytica 206(2):513–521. CrossRefGoogle Scholar
  9. Gebremeskel S, Garcia-oliveira AL, Menkir A, Adetimirin V, Gedil M (2018) Effectiveness of predictive markers for marker assisted selection of pro-vitamin A carotenoids in medium-late maturing maize (Zea mays L.) inbred lines. J Cereal Sci 79:27–34. CrossRefGoogle Scholar
  10. Gupta HS, Hossain F, Muthusamy V (2015) Biofortification of maize: an Indian perspective. Indian J Genet Plant Breed 75(1):1–22. CrossRefGoogle Scholar
  11. Gupta HS, Raman B, Agrawal PK, Mahajan V, Hossain F, Thirunavukkarasu N (2013) Accelerated development of quality protein maize hybrid through marker-assisted introgression of opaque-2 allele. Plant Breed 132(1):77–82. CrossRefGoogle Scholar
  12. Gupta HS, Agrawal PK, Mahajan V, Bisht GS, Kumar A, Verma P et al (2009) Quality protein maize for nutritional security: rapid development of short duration hybrids through molecular marker-assisted breeding. Curr Sci 96:230–237 Google Scholar
  13. Harjes CE, Rocheford TR, Bai L, Brutnell TP, Kandianis CB, Sowinski SG, Stapleton AE, Vallabhaneni R, Williams M, Wurtzel ET, Yan J, Buckler ES (2008) Natural genetic variation in lycopene epsilon cyclase tapped for maize biofortification. Science 319(5861):330–333. CrossRefPubMedPubMedCentralGoogle Scholar
  14. Hernandez HH, Bates LS (1969) A modified method for a rapid tryptophan analysis in maize. CIMMYT, Mexico City, Mexico, Res Bull 13:7 Available at: Google Scholar
  15. Hossain F, Muthusamy V, Pandey N, Vishwakarma AK, Baveja A, Zunjare RU, Thirunavukkarasu N, Saha S, Manjaiah KM, Prasanna BM, Gupta HS (2018) Marker-assisted introgression of opaque2 allele for rapid conversion of elite hybrids into quality protein maize. J Genet 97(1):287–298. CrossRefPubMedGoogle Scholar
  16. Jompuk C, Cheuchart P, Jompuk P, Apisitwanich S (2011) Improved tryptophan content in maize with opaque-2 gene using marker assisted selection (MAS) in backcross and selfing generations. Kasetsart J Nat Sci 45:666–674 Available at: Google Scholar
  17. Kandianis CB, Stevens R, Liu W, Palacios N, Montgomery K, Pixley K, White WS, Rocheford T (2013) Genetic architecture controlling variation in grain carotenoid composition and concentrations in two maize populations. Theor Appl Genet 126(11):2879–2895. CrossRefPubMedPubMedCentralGoogle Scholar
  18. Kennedy G, Nantel G, Shetty P (2003) The scourge of hidden hunger: global dimensions of micronutrient deficiencies. Food Nutr Agric 32:8–16 Available at: Google Scholar
  19. Kumar RS, Kumar B, Kaul J, Karjagi CG, Jat SL, Parihar CM et al (2012) Maize research in India-historical prospective and future challenges. Maize Journal 1:1–6 Available at: Google Scholar
  20. Liu L, Jeffers D, Zhang Y, Ding M, Chen W, Kang MS, Fan X (2015) Introgression of the crtRB1 gene into quality protein maize inbred lines using molecular markers. Mol Breed 35(8):154. CrossRefPubMedPubMedCentralGoogle Scholar
  21. Mertz ET, Bates LS, Nelson OE (1964) Mutant gene that changes protein composition and increases lysine content of maize endosperm. Science 145(3629):279–280. CrossRefPubMedGoogle Scholar
  22. Muthusamy V, Hossain F, Thirunavukkarasu N, Choudhary M, Saha S, Bhat JS, Prasanna BM, Gupta HS (2014) Development of β-carotene rich maize hybrids through marker-assisted introgression of β-carotene hydroxylase allele. PLoS One 9(12):e113583. CrossRefPubMedPubMedCentralGoogle Scholar
  23. Muthusamy V, Hossain F, Thirunavukkarasu N, Saha S, Gupta HS (2015) Allelic variations for lycopene-ɛ-cyclase and β-carotene hydroxylase genes in maize inbreds and their utilization in β-caroteneenrichment programme. Cogent Food Agric 1(1033141).
  24. Neeraja CN, Ravindra Babu V, Ram S, Hossain F, Hariprasanna K, Rajpurohit BS, Prabhakar, Longvah T, Prasad KS, Sandhu JS, Datta SK (2017) Biofortification in cereals : progress and prospects. Curr Sci 113(6):1050–1057. CrossRefGoogle Scholar
  25. Nuss ET, Tanumihardjo SA (2010) Maize: a paramount staple crop in the context of global nutrition. Compr Rev Food Sci Food Saf 9(4):417–436. CrossRefGoogle Scholar
  26. Nyakurwa CS, Gasura E, Mabasa S (2017) Potential for quality protein maize for reducing protein energy undernutrition in maize dependent Sub-Saharan African countries: a review. Afr Crop Sci J 25(4):521–537. CrossRefGoogle Scholar
  27. Rodriguez-Amaya DB (2001) A guide to carotenoid analysis in foods. Washington, ILSI pressGoogle Scholar
  28. Sagare DB, Shetti P, Surender M, Reddy SS, Pradeep T, Anuradha G (2018) Maize: potential crop for provitamin A biofortification. Maydica 63(2):11 Available at: Google Scholar
  29. Selvi DT, Senthil N, Yuvaraj A, Joel AJ, Mahalingam A, Nagarajan P, Vellaikumar S, Srimathi P, Raveendran M, Nepolean T (2014) Assessment of crtRB1 polymorphism associated with increased β-carotene content in maize (Zea mays L.) seeds. Food Biotechnol 28(1):41–49. CrossRefGoogle Scholar
  30. Singh R, Ram L (2014) DNA Aided Introgression of opaque2 allele for development of quality protein maize. Int J Sci Res 3(11): 3006–3011 Available at: googlescholar Google Scholar
  31. Surender M (2015) Conversion of elite non QPM inbreds of maize to QPM using marker assisted selection. Ph.D. In: Thesis. Professor Jayashankar Telangana State Agricultural University, Hyderabad Available at: Google Scholar
  32. Surender M, Shetti P, Sagare DB, Rani CV, Jabeen F, Sudarshan MR et al (2017) Development of QPM version of DHM117 maize hybrid using marker assisted selection. Int J Curr Microbiol Appl Sci 6(10):3275–3289. CrossRefGoogle Scholar
  33. Suwarno WB, Pixley KV, Palacios-Rojas N, Kaeppler SM, Babu R (2015) Genome-wide association analysis reveals new targets for carotenoid biofortification in maize. Theor Appl Genet 128(5):851–864. CrossRefPubMedPubMedCentralGoogle Scholar
  34. Tanumihardjo SA, Anderson C, Kaufer-Horwitz M, Bode L, Emenaker NJ, Haqq AM, Satia JA, Silver HJ, Stadler DD (2007) Poverty, obesity, and malnutrition: an international perspective recognizing the paradox. J Am Diet Assoc 107(11):1966–1972. CrossRefPubMedGoogle Scholar
  35. Vallabhaneni R, Gallagher CE, Licciardello N, Cuttriss AJ, Quinlan RF, Wurtzel ET (2009) Metabolite sorting of a germplasm collection reveals the hydroxylase3 locus as a new target for maize provitamin A biofortification. Plant Physiol 151(3):1635–1645. CrossRefPubMedPubMedCentralGoogle Scholar
  36. Vasal SK (2000) The quality protein maize story. Food Nutr Bull 21(4):445–450. CrossRefGoogle Scholar
  37. Vignesh M, Hossain F, Nepolean T, Supradip S, Agrawal PK, Guleria SK, et al (2012) Genetic variability for kernel beta-carotene and utilization of crtRB1 3’TE gene for biofortification in maize (Zea mays L.). Indian J Genet Plant Breed 72: 189–194 Available at:
  38. Vivek BS, Krivanek AF, Palacios-Rojas N, Twumasi-Afriyie S, Diallo AO (2008) Breeding quality protein maize (QPM): protocols for developing QPM cultivars. Mexico: CIMMYT Available at:
  39. Wong JC, Lambert RJ, Wurtzel ET, Rocheford TR (2004) QTL and candidate genes phytoene synthase and zeta-carotene desaturase associated with the accumulation of carotenoids in maize. Theor Appl Genet 108(2):349–359. CrossRefPubMedGoogle Scholar
  40. Yan J, Kandianis CB, Harjes CE, Bai L, Kim EH, Yang X, Skinner DJ, Fu Z, Mitchell S, Li Q, Fernandez MGS, Zaharieva M, Babu R, Fu Y, Palacios N, Li J, DellaPenna D, Brutnell T, Buckler ES, Warburton ML, Rocheford T (2010) Rare genetic variation at Zea mays crtRB1 increases beta-carotene in maize grain. Nat Genet 42(4):322–327. CrossRefPubMedGoogle Scholar
  41. Yang R, Yan Z, Wang Q, Li X, Feng F (2018) Marker-assisted backcrossing of lcyE for enhancement of proA in sweet corn. Euphytica 214(8):130. CrossRefGoogle Scholar
  42. Zhang X, Pfeiffer WH, Palacios-Rojas N, Babu R, Bouis H, Wang J (2012) Probability of success of breeding strategies for improving pro-vitamin A content in maize. Theor Appl Genet 125(2):235–246. CrossRefPubMedGoogle Scholar
  43. Zhou Y, Han Y, Li Z, Fu Y, Fu Z, Xu S, Li J, Yan J, Yang X (2012) ZmcrtRB3 encodes a carotenoid hydroxylase that affects the accumulation of α-carotene in maize kernel. J Integr Plant Biol 54(4):260–269. CrossRefPubMedGoogle Scholar
  44. Zunjare RU, Hossain F, Muthusamy V, Baveja A, Chauhan HS, Thirunavukkarasu N, Saha S, Gupta HS (2017) Influence of rare alleles of β-carotene hydroxylase and lycopene epsilon cyclase genes on accumulation of provitamin A carotenoids in maize kernels. Plant Breed 136(6):872–880. CrossRefGoogle Scholar
  45. Zunjare RU, Hossain F, Muthusamy V, Baveja A, Chauhan HS, Bhat JS, Thirunavukkarasu N, Saha S, Gupta HS (2018) Development of biofortified maize hybrids through marker-assisted stacking of β-carotene hydroxylase, lycopene-ε-cyclase and opaque2 genes. Front Plant Sci 9(178).

Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  • Deepti B. Sagare
    • 1
    Email author
  • Prashant Shetti
    • 1
  • M. Surender
    • 1
  • S. S. Reddy
    • 1
  1. 1.Department of Molecular Biology and BiotechnologyPJTSAUHyderabadIndia

Personalised recommendations