d. The primers were designed to anneal close (50-75 bp) to the existing 5'-end to

avoid large overlaps.

 Different sets of primers were designed based on the cDNAs obtained by the study

involving the degenerate primers (I4) and the initial 5'-RACE study (I3 5R and L25R).




Table 5-5. Gene-specific primers used in succeeding RLM-RACE study


GSP ID


Sequence (5'-3')


RACE Start' PCR step


(a) Liver L25R

UGT 50UT1
UGT 50041

UGT 30UT1
UGT 3IbW1

(b) Intestine I4
UGT 50UT2
UGT 50UT2A
UGT 5INN2
UGT 5HOWLL

UGT 30UT2
UGT 30UT2A
UGT 3HItz
UGT 3HOWLL


ATTGGGCATTACAGGTCTCG
CGAGGACGTCTCTGAACGTAACATCC


119 Outer
51 Innel

320 Outer
351 Inner


GATTCCTCAGAGGGTTCTGT
GGGGTCATTCCCAAAGACAT


GCCGTTACAGATACCCTCGT
GTATCGCCACAGAACCCTCT
ACACGAAGGAGCTCAAAGTGAACACG
GCCACTTCATCACTTTGACATTTTCAGG


Outer
Outer
Inmer
Imler

Outer
Outer
Imier
Imier


AATGTCAAAGTGATGAAGTGG
GACATTCCTGAAAATGTCAAA
CCCAAGGCTAAGGTGTTCATC
GACCTCTTAGCACACCCCAAG


(b) Intestine I35R


UGT 50UT3
UGT 50UT3A
UGT 5RIN3
UGT 5HIthk

UGT 30UT3
UGT 30UT3A
UGT 3DIN3
UGT 3INN2A


TGTTAATGACCTTCGGTGTGA
ATGACCTTCGGTGTGAGTTTT
AAACCTAAGAGGTCATTCTGCGGAAGC
ATGGGGACCGGGTGTCTATTTATTACG

TTTCCAGCTAACACTACTTGG
TTACACGTCCTCTAACCGTAA
CCCAAGGCTAAGGTGTTCATC
CCATGGCATCTACGAGGGTAT


Outer
Outer
Innel
Imier

Outer
Outer
Innel
Inner


SStart position from partial DNA sequences obtained so far