Tropical Plant Biology

, Volume 1, Issue 3–4, pp 293–309 | Cite as

Characterization of Insertion Sites in Rainbow Papaya, the First Commercialized Transgenic Fruit Crop

  • Jon Y. Suzuki
  • Savarni Tripathi
  • Gustavo A. Fermín
  • Fuh-Jyh Jan
  • Shaobin Hou
  • Jimmy H. Saw
  • Christine M. Ackerman
  • Qingyi Yu
  • Michael C. Schatz
  • Karen Y. Pitz
  • Marcela Yépes
  • Maureen M. M. Fitch
  • Richard M. Manshardt
  • Jerry L. Slightom
  • Stephen A. Ferreira
  • Steven L. Salzberg
  • Maqsudul Alam
  • Ray Ming
  • Paul H. Moore
  • Dennis Gonsalves
Article

Abstract

Inserts and insert sites in transgenic, papaya ringspot virus (PRSV)-resistant commercial papaya Rainbow and SunUp, were characterized as part of a petition to Japan to allow import of fresh fruit of these cultivars from the U.S. and to provide data for a larger study aimed at understanding the global impact of DNA transformation on whole genome structure. The number and types of inserts were determined by Southern analysis using probes spanning the entire transformation plasmid and their sequences determined from corresponding clones or sequence reads from the whole-genome shotgun (WGS) sequence of SunUp papaya. All the functional transgenes, coding for the PRSV coat protein (CP), neophosphotransferase (nptII) and β-glucuronidase (uidA) were found in a single 9,789 basepair (bp) insert. Only two other inserts, one consisting of a 290 bp nonfunctional fragment of the nptII gene and a 1,533 bp plasmid-derived fragment containing a nonfunctional 222 bp segment of the tetA gene were detected in Rainbow and SunUp. Detection of the same three inserts in samples representing transgenic generations five to eight (R5 to R8) suggests that the three inserts are stably inherited. Five out of the six genomic DNA segments flanking the three inserts were nuclear plastid sequences (nupts). From the biosafety standpoint, no changes to endogenous gene function based on sequence structure of the transformation plasmid DNA insertion sites could be determined and no allergenic or toxic proteins were predicted from analysis of the insertion site and flanking genomic DNA.

Keywords

Biosafety Genetically engineered Papaya ringspot virus Particle bombardment Rainbow papaya SunUp papaya Transgene 

Abbreviations

bp

base pair

CP

coat protein

DSB

double-stranded break

ELISA

enzyme-linked immunosorbent assay

FAO

Food and Agriculture Organization (of the United Nations)

GE

genetically engineered

GUS

β-glucuronidase

IUIS

International Union of Immunological Societies

kb

kilobase pair

MAR

matrix attachment regions

NHEJ

nonhomologous end joining

nupt-DNA

nuclear plastid DNA

nupts

nuclear plastid sequence

numts

nuclear mitochondrial sequence

ORF

open reading frame

PCR

polymerase chain reaction

PDR

pathogen-derived resistance

PTGS

post-transcriptional gene silencing

PRSV

Papaya ringspot virus

SDAP

Structural Database for Allergenic Proteins

T-DNA

transferred DNA

Topo I

Topoisomerase I

Topo II

Topoisomerase II

WGS

whole-genome shotgun

WHO

World Health Organization

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Copyright information

© Springer Science+Business Media, LLC 2008

Authors and Affiliations

  • Jon Y. Suzuki
    • 1
  • Savarni Tripathi
    • 1
    • 7
  • Gustavo A. Fermín
    • 2
  • Fuh-Jyh Jan
    • 3
  • Shaobin Hou
    • 4
  • Jimmy H. Saw
    • 4
    • 11
  • Christine M. Ackerman
    • 5
  • Qingyi Yu
    • 5
  • Michael C. Schatz
    • 6
  • Karen Y. Pitz
    • 7
  • Marcela Yépes
    • 8
  • Maureen M. M. Fitch
    • 1
  • Richard M. Manshardt
    • 9
  • Jerry L. Slightom
    • 10
  • Stephen A. Ferreira
    • 7
  • Steven L. Salzberg
    • 6
  • Maqsudul Alam
    • 4
    • 11
  • Ray Ming
    • 5
    • 12
  • Paul H. Moore
    • 1
  • Dennis Gonsalves
    • 1
  1. 1.USDA-ARS Pacific Basin Agricultural Research CenterHiloUSA
  2. 2.Centro Jardín BotánicoUniversidad de los AndesMéridaVenezuela
  3. 3.Department of Plant PathologyNational Chung Hsing UniversityTaichung, TaiwanRepublic of China
  4. 4.Advanced Studies in Genomics, Proteomics and BioinformaticsUniversity of HawaiiHonoluluUSA
  5. 5.Hawaii Agricultural Research CenterHonoluluUSA
  6. 6.Center for Bioinformatics and Computational BiologyUniversity of MarylandCollege ParkUSA
  7. 7.Plant and Environmental Protection Sciences, College of Tropical Agriculture and Human ResourcesUniversity of HawaiiHonoluluUSA
  8. 8.Department of Plant PathologyCornell UniversityGenevaUSA
  9. 9.Department of Tropical Plant and Soil Sciences, College of Tropical Agriculture and Human ResourcesUniversity of HawaiiHonoluluUSA
  10. 10.AureoGen BiosciencesKalamazooUSA
  11. 11.Department of MicrobiologyUniversity of HawaiiHonoluluUSA
  12. 12.Department of Plant BiologyUniversity of Illinois at Urbana-ChampaignUrbanaUSA

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