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Plant Growth Regulation

, Volume 88, Issue 1, pp 61–71 | Cite as

Improvement of post-harvest fruit characteristics in tomato by fruit-specific over-expression of oat arginine decarboxylase gene

  • Aarti Gupta
  • Roopali Pandey
  • Ranjita Sinha
  • Anuj Chowdhary
  • Ram Krishna Pal
  • Manchikatla Venkat RajamEmail author
Original paper
  • 71 Downloads

Abstract

Over-ripening is the key contributor to post-harvest loss of perishable fruits and is also one of the greatest threats to the growers. Delayed ripening of tomato fruits has been suggested as a possible solution to such post-harvest losses. To overcome post-harvest losses, we engineered tomato fruits to accumulate polyamines (PAs) by over-expression of arginine decarboxylase (ADC) gene of PA biosynthetic pathway. The AsADC transgenic fruits contained elevated levels of putrescine, spermidine and spermine content in comparison to the fruits from untransformed plants. In addition, the expression of several ripening-associated genes was also altered. The transgenic fruits displayed a better shelf-life due to decrease in ethylene levels and respiratory activity. The transgenic tomatoes also exhibited improved key processing features along with the elevated levels of lycopene and vitaminC. Our study highlights that the fortification of PAs is a promising approach for the improvement of post-harvest fruit characteristics together with a high quality of produce.

Keywords

Polyamines Tomato Arginine decarboxylase Fruit-specific promoter Transgenic plants Fruit quality 

Notes

Acknowledgements

We thank Dr. R. L. Malmberg (University of Georgia, USA) for providing the AsADC cDNA (from Avena sativa) and Dr. V. S. Reddy (International Centre for Genetic Engineering and Biotechnology, New Delhi) for sharing the 2A11 tomato promoter. This work was supported by grants from the Department of Biotechnology (Govt. of India), New Delhi (Grant No. BT/PR/2990/Agr/16/232/2002 and BT/PR8657/PBD/16/738/2007), University Grants Commission for Special Assistance Programme (DRS-III) and Department of Science and Technology (DST) for FIST (Level 2) programme and DU-DST PURSE Phase II grant. AG and RP acknowledge the Council of Scientific and Industrial Research, New Delhi and RS acknowledgesthe Department of Biotechnology for the research fellowships.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

10725_2019_488_MOESM1_ESM.jpg (43 kb)
Fig. S1. Fruit-specific over-expression of AsADC cDNA in transformed tomato plants. (A) Transfer (T)-DNA map of the pBin2A11ADC binary vector is shown here. LB, left border of T-DNA; NOST, terminato of nopaline synthase gener; NPT-II, neomycin phosphotransferase-II gene; NOSP, nos promoter; 2A11P, 2A11 promoter; OCST, terminator of octopine synthase gene; RB, right border of T-DNA. Arrow heads denote the direction of promoter activity. (B) RNA blot image depicting transgene expression in ripening fruits from UT and AsADC51 transgenic plant is presented here. RNA blot analysis was carried out using radio-labeled AsADCgene fragment to probe transgene transcript accumulation in T3 generation fruits. Radio-labeled SlACTIN gene probe was used to ascertain ACTIN gene levels as an internal control. MG, mature green; BR, breaker; PR, pinkred; RR, red ripe stages of fruit ripening. Supplementary material 1 (JPG 43 KB)
10725_2019_488_MOESM2_ESM.jpg (140 kb)
Fig. S2. Molecular characterization of AsADC transgenic tomato plants. (A) Gel image showing PCR with primers specific to NPT-II gene in primary transformants is shown here. L, 1 kb ladder; PC, Plasmid DNA; UT, DNA from UT control; AsADC33-93, DNA from AsADC transgenic tomato lines. (B) Image showing Southern blot analysis of AsADC transgenic plants using radio-labeledNPT-II gene probe detailing gene integration and copy number is presented here. UT, DNA from UT control; AsADC33-92, DNA from different transgenic lines digested with XbaI enzyme. (C) A flow-diagram showing scheme adopted for analyzing the segregation pattern of the transgene from T1 to T3 generation. Leaf from 100 T1 seedlings (named as e. g., AsADC51-1 to AsADC51-100) of each positive T0 plant (e.g., AsADC51) was screened by kanamycin resistant assay and 40 positive segregants were further confirmed by AsADC specific PCR. 10 positive segregants were taken forward to T2 generation. 100 T2 seedlings (further named as e.g., AsADC51-1.1 to AsADC51-1.100.) from each of these 10 such parents were further screened by kanamycin resistance assay and confirmed by PCR. The leaves of T1 transgenic line AsADC51. (D) Photograph showing segregation of the ADC transgene on kanamycin (50 mg/L) amended MS medium, which was based on the bleaching of leaves are shown here. The UT leaves, ‘c’ showed complete bleaching on antibiotic medium. E1 and E2 show gel images depicting PCR analysis of AsADC51 T2 and T3 progenies with primers specific to AsADC gene showing segregation pattern of the transgene are presented here. L;-1 kb DNA ladder, PC; Plasmid DNA, NC; DNA from UT control, 1-12; DNA from different progenies of AsADC51 line. Supplementary material 2 (JPG 141 KB)
10725_2019_488_MOESM3_ESM.jpg (48 kb)
Fig. S3. Expression analysis of AsADC transgene in different organs of the transgenic tomato plant. Gel image showing semi-quantitative-RT-PCR based transgene expression in different organs from UT and AsADC51 transgenic plant is presented here. Semi-quantitative RT-PCR analysis for ACTINgene was performed as an internal control. L; 1 kb ladder,LF; leaf, FLB;flower bud, IG; immature green, MG; mature green, BR; breaker. Supplementary material 3 (JPG 49 KB)
10725_2019_488_MOESM4_ESM.doc (51 kb)
Supplementary material 4 (DOC 51 KB)

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Copyright information

© Springer Nature B.V. 2019

Authors and Affiliations

  • Aarti Gupta
    • 1
    • 3
  • Roopali Pandey
    • 1
  • Ranjita Sinha
    • 1
  • Anuj Chowdhary
    • 2
  • Ram Krishna Pal
    • 2
  • Manchikatla Venkat Rajam
    • 1
    Email author
  1. 1.Plant Polyamine, Transgenic and RNAi Research Laboratory, Department of GeneticsUniversity of Delhi South CampusNew DelhiIndia
  2. 2.Division of Post-Harvest TechnologyIndian Agricultural Research InstituteNew DelhiIndia
  3. 3.Institute of Plant GeneticsPolish Academy of SciencesPoznanPoland

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