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Chapter 3 on AMPs. These molecules act as endogenous antibiotics and as such are considered to be a key element of innate immunity (Bulet et al., 1999). More than 1000 AMPs have now been discovered in plants, vertebrates, and invertebrates. They are mostly cationic and amphipathic molecules and show great diversity in terms of structural features as well as biological properties and functions. Due to their small size, amphipathic structure and cationic character they can rapidly diffuse to the point of infection (Brogden, 2005), a mechanism that presumably makes it easier to circumvent microbial resistance (Bax et al., 2000). AMPs can kill bacteria in micromolar range, promptly synthesized at low metabolic cost, easily stored in large amounts and readily available shortly after an infection, to rapidly kill a broad range of microbes (Hancock, 1997, 2001; Prenner et al., 1999). Many antibacterial peptides show a remarkable specificity for prokaryotes with low toxicity for eukaryotic cells; a phenomenon which has favored their investigation and exploitation as potential new antibiotics (Zasloff, 1992). The discovery of AMPs in crustaceans provides new clues for fundamental understanding of crustacean immunity. The studies on the shrimp penaeidins have largely contributed to this knowledge as they are the first AMPs fully characterized in a crustacean and for which expression studies were performed. Soon after their discovery and initial characterization they have been subjects of extensive studies at structural and functional level. This chapter represents the first report of AMPs from Indian white shrimp, F. indicus. The reported AMPs belonged to the class of ALFs, crustins and penaeidins; the three major groups of AMPs characterized from penaeid shrimps. ALF is a small basic protein which was initially isolated and characterized from haemocytes of the horseshoe crab L. polyphemus (Aketagawa et al., 1986; Muta et al., 1987). L. polyphemus ALF binds LPS and has a strong antibacterial activity, particularly on the growth of Gram- Molecular Characterization and Gene Expression Profiling of Antimicrobial Peptides in Penaeid Shrimps 200
Chapter 3 negative bacteria. In shrimp, cDNA clones homologous to the horseshoe crab ALFs were initially identified in haemocytes of P. monodon and L. setiferus by expressed sequence tag (EST) analysis (Gross et al., 2001; Supungul et al., 2004). More recently, a similar approach has been implemented to identify ALF homologues in other shrimp species including the Chinese shrimp F. chinensis (Liu et al., 2005) and the kuruma shrimp M. japonicus (Nagoshi et al., 2006). Antimicrobial assays using recombinant P. monodon ALF have shown strong activity against multiple gram-positive and gram-negative bacteria and filamentous fungi (Somboonwiwat et al., 2005). In the present study two isoforms of ALFs could be characterized. The complete coding sequence of the ALF-2 could also be obtained in the present study. In the present study two putative ALFs could be characterized from the haemocytes of the Indian white shrimp, F. indicus. ALF-1 was found to be a new isoform of ALF from F. indicus. However, only a partial sequence of ALF-1 could be characterized. Complete coding sequence of the ALF-2 could be characterized from the haemocytes of F. indicus. The ALF-2 sequence shared 98% similarity with another ALF recently reported from the same species. Results of BLAST, multiple alignments and phylogenetic study revealed both ALF-1 and ALF-2 to be closely related isoforms of the ALF family. ALFs characterized from F. indicus haemocytes were found to be more related to other ALFs of Fenneropenaeus sp. than to Litopenaeus, Farfantepenaeus or Penaeus sp. The molecular weight, pI and amino acid composition predicted out for ALF-2 were in agreement with the results of Beale et al. (2008) and Imjongjirak et al. (2007). Peptide model of the partial ALF-1 created using SWISS-MODEL server, was found to consist of both alpha-helical structure and β-sheets (Fig.3.3). Four β-sheets and one α-helix could be detected in the ALF-1 of F. indicus. As per Hoess et al. (1993) and Beale et al. (2008), the ALF molecules should consist of a single domain with Molecular Characterization and Gene Expression Profiling of Antimicrobial Peptides in Penaeid Shrimps 201
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Molecular Characterization and Gene
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Declaration I hereby do declare tha
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Acknowledgements This work was carr
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My Seniors, Dr. Annies Joseph, Dr.
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Contents Title Page Nos. 1 General
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in response to WSSV challenge 252-2
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α-2M Alpha-2-Macroglobulin ALF Ant
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ppA Prophenoloxidase-Activating Enz
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1.1 Introduction Chapter 1 Aquacult
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Chapter 1 the giant tiger shrimp (P
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Common names: Chapter 1 Giant tiger
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1.3.1. Diseases Chapter 1 Diseases
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Lymphoidal parvo-like virus (LPV) B
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Chapter 1 and protective occlusion
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Chapter 1 small brown masses in the
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1.4.6. Natural epidemics Chapter 1
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Chapter 1 can differentiate WSSV-in
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1.5.1.1. Probiotics Chapter 1 Alter
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Chapter 1 tried in P. monodon, resu
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Chapter 1 The first and essential i
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Chapter 1 1998). The innate immunit
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1.6.1 Haemocytes Chapter 1 The haem
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Chapter 1 In crustaceans, the sheet
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Chapter 1 Theopold et al., 2002). C
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Chapter 1 kill/remove the invading
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Chapter 1 forming complexes with a
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1.7 Antimicrobial Peptides (AMPs) 1
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Chapter 1 the response is very fast
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Chapter 1 Hirsch, 1947). While they
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Chapter 1 AMPs to various degrees u
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1.7.5 Biological activity Chapter 1
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Chapter 1 including the microbes as
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Tachyplesin I Chapter 1 Crustacea P
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Anionic and cationic peptides that
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Anionic and Cationic AMPs with cyst
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Chapter 1 Fig. 1.7. Structural clas
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Chapter 1 Group III: Linear peptide
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Chapter 1 through regular processes
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Chapter 1 ultimately signal to tran
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Chapter 1 divalent polyanionic cati
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Chapter 1 type of transmembrane por
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Chapter 1 other intracellular targe
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Chapter 1 Apidaecin, a short, proli
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Chapter 1 Hultmark, 1987; Gennaro a
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Chapter 1 quite opposite characteri
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Chapter 1 depletion of mitochondria
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Chapter 1 conformations adopted by
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Chapter 1 receptor may be a potenti
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Chapter 1 exposure to their targets
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Chapter 1 generating a rich spectru
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Chapter 1 by enkelytin (Goumon et a
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Lactoferricin Cow Mastitis control
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Chapter 1 trial, in which peptide T
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Chapter 1 a too limited spectrum to
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Chapter 1 number of research groups
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CHAPTER-2 Molecular Characterizatio
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Chapter 2 interaction with negative
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Chapter 2 inhibit the endotoxin or
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Chapter 2 structure be modified to
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Chapter 2 and the region might be l
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Chapter 2 been made of the spectra
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Chapter 2 their discovery and initi
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Chapter 2 terminal pyroglutamic aci
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Chapter 2 One can assume that the p
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Chapter 2 by degranulation into the
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2.2. Materials and Methods 2.2.1. E
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Chapter 2 washed by adding 1 ml 75
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Chapter 2 entry of recombinant DNA
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Chapter 2 and the deduced amino aci
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Chapter 2 Fenneropenaeus, Litopenae
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2.3.1.3. Crustin-1 (GQ334395) Chapt
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2.3.1.3.5. Phylogenetic analysis of
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Chapter 2 sequence also showed high
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Chapter 2 chinensis and F. indicus
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Chapter 2 monodon and Group III of
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Chapter 2 Tiger shrimp (P. monodon)
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Chapter 2 ALF-1 was a partial seque
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Chapter 2 As mentioned before, the
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Chapter 2 Munoz et al., 2004). The
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Table 2.1. Primers used for the stu
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Table 2.4. BLASTn analysis of ALF-2
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Table 2.8. BLASTn analysis of Crust
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Table 2.12. BLASTn analysis of Pena
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Page 177 and 178: Chapter 2 Fig.2.20. Signal peptide
Page 179 and 180: Chapter 2 Fig.2.24 Multiple alignme
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Page 183 and 184: Chapter 2 Fig. 2.28. Nucleotide and
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Page 197 and 198: Chapter 2 LITOPENAEUS SP. FENNEROPE
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Page 201 and 202: Chapter 2 FENNEROPENAEUS SP. FARFAN
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Page 211 and 212: Chapter 3 random coiled structure t
Page 213 and 214: 3.3.1.3.5. Phylogenetic analysis of
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Page 217: Chapter 3 Multiple alignments were
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Page 223 and 224: Chapter 3 horseshoe crab big defens
Page 225 and 226: Table 3.4. BLASTn analysis of ALF-2
Page 227 and 228: Table.3.8. BLASTn analysis of Fi-pe
Page 229 and 230: Chapter 3 Fig.3.2. Nucleotide and a
Page 231 and 232: Chapter 3 Fig.3.5. Multiple alignme
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Page 237 and 238: Chapter 3 Fig.3.14. A bootstrapped
Page 243 and 244: SHRIMPS KIND CRAB LOBSTERS CRABS Ch
Page 247 and 248: Chapter 3 Fig. 3.28. Multiple align
Page 249 and 250: PENAEIDIN-2 Chapter 3 PENAEIDIN- 3
Page 251 and 252: Fig.3.31. Nucleotide and amino acid
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Page 257 and 258: 4.1. Introduction Chapter 4 The aqu
Page 259 and 260: Chapter 4 appearance and have major
Page 261 and 262: Chapter 4 the tissues has recently
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Chapter 4 for understanding gene ex
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Chapter 4 4.3.2. Expression profile
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Expression profile of endonuclease
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Chapter 4 bacterial and fungal infe
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Chapter 4 measuring TSV and YHV loa
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Chapter 4 Of particular interest he
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Chapter 4 large distribution and ab
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Chapter 4 WSSV genes viz. endonucle
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Table 4.2. Primers used for the stu
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(A) (B) B 28 1 2 3 4 5 6 7 8 9 10 C
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(A) (B) B 28 1 2 3 4 5 6 7 8 9 10 C
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(A) (B) B 28 1 2 3 4 5 6 7 8 9 10 C
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(A) (B) B 28 1 2 3 4 5 6 7 8 9 10 C
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(A) (B) B 28 1 2 3 4 5 6 7 8 9 10 F
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(A) (B) B 28 1 2 3 4 5 6 7 8 9 10 C
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(A) (B) B 28 1 2 3 4 5 6 7 8 9 10 C
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(A) (B) B 28 1 2 3 4 5 6 7 8 9 10 C
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CHAPTER-5 Expression Profile of Ant
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Chapter 5 1994). The emergence of b
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Chapter 5 (Bovill et al., 2001). In
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5.2.2. Test diets Chapter 5 Four ex
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5.2.2.4. Preparation of Glucan inco
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Chapter 5 genes (latency related ge
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Chapter 5 On WSSV challenge, crusti
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Chapter 5 5.3.4. Expression profile
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5.3.4.5. Expression profile of pena
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Chapter 5 must firstly rest on a so
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Chapter 5 For the present study two
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Chapter 5 ameobocytes of two marine
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Chapter 5 variation in expression o
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Chapter 5 It has been previously sh
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Chapter 5 manifested in terms of gr
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(A) (B) (C) C CHY CSY CHG CSG C CHY
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(A) (B) -VE CTRL +VE CTRL CHG CSG C
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(A) PRE-CHALLENGE (B) POST-CHALLENG
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(A) PRE-CHALLENGE (B) (C) (D) CONTR
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(C) (D) (A) PRE-CHALLENGE (B) CONTR
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(A) (B) G M HP HR I Chapter 5 Fig.
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Chapter 5 Fig. 5.26. Post challenge
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6.1. Introduction Chapter 6 Prophyl
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Chapter 6 production of inhibitory
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6.2. Materials and methods 6.2.1. E
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Chapter 6 tissue cDNA was performed
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Chapter 6 gene was supported by Bac
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Chapter 6 treated group of shrimps.
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Chapter 6 the gene was found to be
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Chapter 6 found to support minimum
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Chapter 6 is induced upon bacterial
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Chapter 6 all the target genes, exc
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Chapter 6 Detailed tissue-wise anal
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Chapter 6 Gills and intestine was f
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(A) (B) Fig. 6.1. Experimental diet
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(A) (B) (C) Fig. 6.3. Expression pr
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(A) (B) (C) C B M BM C B M BM Fig.
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(A) (B) (C) Fig. 6.7. Expression pr
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(A) (B) (C) Chapter 6 Fig. 6.9. Exp
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(A) (B) -VE CTRL +VE CTRL B M BM Ch
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(A) (C) (D) PRE-CHALLENGE POST-CHAL
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(A) (B) G M HP HR I Chapter 6 Fig.
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Chapter 6 Fig. 6.27. Post challenge
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7.1. Summary Chapter 7 Tropical shr
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Salient findings Chapter 7 � From
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Chapter 7 � Tissue-wise expressio
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Chapter 7 application in shrimp cul
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References Aboudy, Y., Mendelson, E
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References Anggraeni, M.S., Owens,
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References Bals, R., Wang, X., Zasl
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References derived from the N-termi
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References Breukink, E., de Kruijff
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References Burgents, J.E., Burnett,
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References 1,3-β-D-glucan-binding
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References Chen, J.Y., Chuang, H.,
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References Cole, A.M., Hong, T., Bo
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References Dathe, M., Wieprecht, T.
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References Diamond, G., Zasloff, M.
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References Faber, C., Stallmann, H.
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References Flint, S.J., Enquist, L.
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References Gennaro, R., Zanetti, M.
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References Gudmundsson, G.H., Lidho
Page 449 and 450:
References Harwig, S.S., Swiderek,
Page 451 and 452:
References Hirakura, Y., Kobayashi,
Page 453 and 454:
References Huang, C.C., Song, Y.L.
Page 455 and 456:
References Itami, T., Takahashi, Y.
Page 457 and 458:
References Jiravanichpaisal, P. Whi
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References Kang, J.H., Shin, S.Y.,
Page 461 and 462:
References Kono, T., Savan, R., Sak
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References rich peptide derived fro
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References Li, L., Wang, J.X., Zhao
Page 467 and 468:
References Liu, C.H., Cheng, W., Ku
Page 469 and 470:
References Lorenzon, S., de Guarrin
Page 471 and 472:
References Marone, M., Mozzetti, S.
Page 473 and 474:
References Medvinsky, A., Dzierzak,
Page 475 and 476:
References Moriarty, D.J.W. 1996. M
Page 477 and 478:
References antimicrobial peptide fr
Page 479 and 480:
References Otta, S.K., Karunasagar,
Page 481 and 482:
References simplex virus has heptad
Page 483 and 484:
References infected shrimp. Thesis
Page 485 and 486:
References Litopenaeus vannamei cha
Page 487 and 488:
References Roth, B.L., Poot, M., Yu
Page 489 and 490:
References Sathish, S., Musthaq, S.
Page 491 and 492:
References a proline-arginine-rich
Page 493 and 494:
References Smith, V.J., Fernandes,
Page 495 and 496:
References Song, Y.L., Cheng, W., W
Page 497 and 498:
References experimental system. In:
Page 499 and 500:
References the black tiger shrimp P
Page 501 and 502:
References Tapay, L.M., Cesar, E.,
Page 503 and 504:
References Touhy, K.M., Probert, H.
Page 505 and 506:
References van Hulten, M.C., Reijns
Page 507 and 508:
References Vives, R.R., Imberty, A.
Page 509 and 510:
References Wang, Y.C., Chang, P.S.,
Page 511 and 512:
References Williams, M.J., Rodrigue
Page 513 and 514:
References Wu, W., Zong, R., Xu, J.
Page 515 and 516:
References Yodmuang, S., Tirasophon
Page 517 and 518:
References Zhan, W.B., Wang, Y.H. 1
Page 519 and 520:
Penaeus monodon anti-lipopolysaccha
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Penaeus monodon crustin-like antimi
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Penaeus monodon crustin-like antimi
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Penaeus monodon penaeidin-like anti
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Penaeus monodon elongation factor m
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Fenneropenaeus indicus anti-lipopol
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Fenneropenaeus indicus penaeidin-li
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Fenneropenaeus indicus beta-actin m
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PUBLICATIONS
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Short sequence report Molecular cha
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Fig. 2. Multiple alignment of nucle
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220 The phylogenetic relationships
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Table 1 Rearing conditions and wate
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5 ′ cttgtggttgacaatggctccg3
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Expression of WSSV genes in the hae
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Fig. 4. Expression profile of crust
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S.P. Antony et al. / Immunobiology
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